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Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X Plan-NeoFluar

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  • Charles Guevara
       Thank you so much, Rich, for this practicle/hands on microscopy thread.     Rolf, Randall, David have clarified performance for visual, vrs. image
    Message 1 of 21 , Nov 1, 2011
         Thank you so much, Rich, for this practicle/hands on microscopy thread.
       
        Rolf, Randall, David have clarified performance for visual, vrs. image capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction collar for coverslip/water colum variations?.
       
         Rolf noted that the specimen context: (wetmount-pondlife slides with coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf noted that this context renders: 'Plan objectives utility as often over rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your bloodwork observations,Rich,...try for oil-immersion 100X objectives (even though you do not like to muddle with oiled-optics...as a 'fall back...Rolf suggested : water immersion/LOMO objectives.).
       
         I have : NA 1.25 condensors on both my work-horse stands....I have a Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have Nikon CFW 10X occulars with this setup.
       
       
          I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars with this setup.  
       
         Please...this is not a 'high-jack', of Rich's thread...please comment to Rich (...err, to all)...although we rarely do it...why is not: 'oil-bridgeing the substage-condensor not at least mentioned in this discussion with Rich pondering objectives for his use( okay...Rich already said he is not keen on imerrsion oil useage)?
       
         Please...as I have a variety of occulars...( okay...I think Rich already said he is useing:'10X occulars'.)...please comment on how one need concern a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X PlanApo?
       
       
          Do occulars even matter in Rich-guys setup?  With the very common: NA 1.25 substage condensors...we often do not 'oil-bridge condensor to slide'..does this factor in to Rich'es 'application Difference'?    thanks for all this practicle/ applied microscopy, charlie guevara     (PS)  in hobby astronomy...Apo refractors are costly, their proud owners seek out planetary observing agendas for crisp/'tack-sharp'  color observations.   Deep-sky (beyond our Suns family of interacting systems)...deep sky observers always 'chant': so little color differences in 'deep sky objects'...no need to worry about: 'purple fringes on bright stars...go 'achro'...leave 'apos' for when you win the lottery.

      From: Randall Buck <rbuck@...>
      To: Microscope@yahoogroups.com
      Sent: Sunday, October 30, 2011 8:31 PM
      Subject: RE: [Microscope] Application Difference?? 100X PlanApo vs, 63X Plan-NeoFluar

      Hi Rich,

      I will have to go with Rolf and David.

      Think of a high NA apo objective as a race car.  Sure, you can take it to
      the store to get bread and milk but since it is so finely tuned to run
      within such a narrow range of specifications, you may have to give it a tune
      up before you go.  In fact, you may spend more time tuning it than it takes
      to perform the errand.

      High NA Dry Objectives:
      Glare arises from reflections between the top of the coverslip and the front
      lens.  Higher power dry objectives have high NAs and so have small working
      distances; this leads to a greater possibility for contrast reducing
      reflection glare of this origin.  Add to that, the fact that a 63X dry will
      be very sensitive to coverslip thickness and you have a tricky lens to use.

      In general, an oil immersion lens will not be nearly as sensitive to cover
      slip thickness.  In addition, reflection glare is virtually eliminated with
      oil immersion objectives (assuming the oil index is quite close to both the
      front lens and the coverslip index of refraction).

      If your everyday condenser can be oiled (for a homogenous immersion
      condition), I doubt you would notice a difference between it and your
      aplanatic achromatic condenser (always oiled) until you get past NA 1.3 for
      your objective.  So maybe it is a lucky break, missing that planapo?

      For Example:
      I have a Leitz NPL Fluotar, 100X, 1.32, oil that yields very nice images
      with a 0.9 NA dry condenser.  It is very easy to use and I didn't have to
      hold up a convenience store just to pay for it.  If I really need a bit more
      from it, then I can switch to an aa condenser.  With a NA 1.4 objective, the
      aa condenser might be the perfect choice -- I don't know, I don't have a 1.4
      NA objective.

      Randall




      -----Original Message-----
      From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
      Behalf Of rich_guy_looker
      Sent: Tuesday, October 18, 2011 11:56 AM
      To: Microscope@yahoogroups.com
      Subject: [Microscope] Application Difference?? 100X PlanApo vs, 63X
      Plan-NeoFluar


      I just saw on Ebay; a 100X PlanApo [oil],
      with an 0.8 to 1.25 NA [iris]
      go for about $400.00

      I wanted that lens,
      but my limit was $325.00

      I already have a [dry] 63x Plan-Neofluar
      with 0.9 NA and a ach/apl condenser,
      so I actually could go up to 900X magnification.

      I'm looking at human circulatory components
      (5 to 20 micrometers in size).

      QUESTION:
      Should I be "upset and bent-out-of-shape"
      because there would have been a big difference
      in the image quality between the 2 lenses above??

      My nerves are absolutely frazzeled
      right now.

      Rich






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    • Randall Buck
      Hi, The more I think about the subject, the more I come to believe that John W. (ajohn) has condensed the whole subject into a very succinct statement:
      Message 2 of 21 , Nov 2, 2011
        Hi,

        The more I think about the subject, the more I come to believe that John W.
        (ajohn) has condensed the whole subject into a very succinct statement:
        (Paraphrasing), Think of Very high NA objectives as though they are designed
        for one thing only, viewing something that is attached to the underside of a
        precision coverslip (one that meets the design specifications of the
        objective in hand).

        You certainly don't want to be looking through anything other than immersion
        oil * and the coverslip otherwise you will introduce an amount of spherical
        aberration that will reduce the performance your expensive objective to that
        of an objective for which you could have paid much less. Even mountant,
        unless it is more of a filler than an intervening layer may require a
        thinner coverslip to compensate for any added thickness.

        Also, remember that in order to take full advantage of the high NA of the
        objective, you will need a high NA condenser. Since the condenser is
        operating at high NA it requires improved optical correction to deliver a
        precisely imaged beam to the subject plane. This is why the aplanatic
        achromat was developed. Of course, it must be oiled to the bottom of the
        slide, in fact, it will generally be impossible to align it of Kohler
        without oiling. In addition, the iris should be used wide open, otherwise
        your expensive 1.4NA aa condenser will be doing the job of a much less
        expensive (lower NA) condenser and taking the performance of your High NA
        objective down with it.

        This means that the perceived contrast improvement ordinarily gained by
        reducing the condenser iris to 2/3 cannot be employed when the maximum
        resolution of a high NA objective is the goal. In turn, this means that
        the specimen itself must be of intrinsically high contrast (via staining,
        etc) to be seen at all.

        That is not to say that pursuing ultimate resolution isn't worth while or
        fascinating in and of itself but it is a very precise endeavor.

        NOTE:

        * Older objectives were designed for appropriately aged cedar oil with each
        lens manufacturer providing their own optimum formulation. The index of
        refraction of cedar oil increases with aging. Newer (does anyone know the
        approximate transition date?) objectives were designed for Shillaber's (now
        Cargille) non-drying synthetic oils, with a (sodium "D" line index of
        refraction of 1.5150 at 25 deg C).


        Randall





        -----Original Message-----
        From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
        Behalf Of Charles Guevara
        Sent: Tuesday, November 01, 2011 7:53 PM
        To: Microscope@yahoogroups.com
        Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
        Plan-NeoFluar


           Thank you so much, Rich, for this practicle/hands on microscopy thread.
         
          Rolf, Randall, David have clarified performance for visual, vrs. image
        capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
        Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction collar
        for coverslip/water colum variations?.
         
           Rolf noted that the specimen context: (wetmount-pondlife slides with
        coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
        noted that this context renders: 'Plan objectives utility as often over
        rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
        bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
        though you do not like to muddle with oiled-optics...as a 'fall back...Rolf
        suggested : water immersion/LOMO objectives.).
         
           I have : NA 1.25 condensors on both my work-horse stands....I have a
        Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
        objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
        Nikon CFW 10X occulars with this setup.
         
         
            I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
        with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars with
        this setup.  
         
           Please...this is not a 'high-jack', of Rich's thread...please comment to
        Rich (...err, to all)...although we rarely do it...why is not:
        'oil-bridgeing the substage-condensor not at least mentioned in this
        discussion with Rich pondering objectives for his use( okay...Rich already
        said he is not keen on imerrsion oil useage)?
         
           Please...as I have a variety of occulars...( okay...I think Rich already
        said he is useing:'10X occulars'.)...please comment on how one need concern
        a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
        PlanApo?
         
         
            Do occulars even matter in Rich-guys setup?  With the very common: NA
        1.25 substage condensors...we often do not 'oil-bridge condensor to
        slide'..does this factor in to Rich'es 'application Difference'?    thanks
        for all this practicle/ applied microscopy, charlie guevara     (PS)  in
        hobby astronomy...Apo refractors are costly, their proud owners seek out
        planetary observing agendas for crisp/'tack-sharp'  color observations.  
        Deep-sky (beyond our Suns family of interacting systems)...deep sky
        observers always 'chant': so little color differences in 'deep sky
        objects'...no need to worry about: 'purple fringes on bright stars...go
        'achro'...leave 'apos' for when you win the lottery.
      • Selwyn St Leger
        HiRandall, Those are interesting observations and well put. One point you raise merits further discussion. That is whether or not the condenser diaphragm
        Message 3 of 21 , Nov 3, 2011
          HiRandall,

          Those are interesting observations and well put.

          One point you raise merits further discussion. That is whether or not the condenser diaphragm should be reduced below NA 1.4 when employing a 1.4 NA objective.

          With geometric optics the condenser diaphragm is conjugate with (in focus at) the back focal plane of the objective. Thus, from this way of considering light transmission its effect is exactly the same as an equivalent stop in the back focal plane. However, as the image is formed from diffracted rays off the specimen some of these pass beyond the geometric light cone, impinge on the periphery of the objective to contribute to interference at the back focal plane.

          Thus, stopping down the condenser a little does not have a large effect on resolution. Also, apochromatic objectives (which they must be to attain 1.4 NA) because of their corrections require less stopping down of the condenser than achromats to achieve good contrast.

          Nevertheless, your broader point has great merit. Attempting to get maximum resolution is not often, in contexts such as mine, worth the candle. Very high resolution has to be accompanied by the facility to interpret what has been resolved. That's fine with test diatoms but looking, say, at living cytoplasm in a protist there is not much point in being able to perceive a few more anonymous granules.

          Selwyn


          --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@...> wrote:
          >
          >
          > Hi,
          >
          > The more I think about the subject, the more I come to believe that John W.
          > (ajohn) has condensed the whole subject into a very succinct statement:
          > (Paraphrasing), Think of Very high NA objectives as though they are designed
          > for one thing only, viewing something that is attached to the underside of a
          > precision coverslip (one that meets the design specifications of the
          > objective in hand).
          >
          > You certainly don't want to be looking through anything other than immersion
          > oil * and the coverslip otherwise you will introduce an amount of spherical
          > aberration that will reduce the performance your expensive objective to that
          > of an objective for which you could have paid much less. Even mountant,
          > unless it is more of a filler than an intervening layer may require a
          > thinner coverslip to compensate for any added thickness.
          >
          > Also, remember that in order to take full advantage of the high NA of the
          > objective, you will need a high NA condenser. Since the condenser is
          > operating at high NA it requires improved optical correction to deliver a
          > precisely imaged beam to the subject plane. This is why the aplanatic
          > achromat was developed. Of course, it must be oiled to the bottom of the
          > slide, in fact, it will generally be impossible to align it of Kohler
          > without oiling. In addition, the iris should be used wide open, otherwise
          > your expensive 1.4NA aa condenser will be doing the job of a much less
          > expensive (lower NA) condenser and taking the performance of your High NA
          > objective down with it.
          >
          > This means that the perceived contrast improvement ordinarily gained by
          > reducing the condenser iris to 2/3 cannot be employed when the maximum
          > resolution of a high NA objective is the goal. In turn, this means that
          > the specimen itself must be of intrinsically high contrast (via staining,
          > etc) to be seen at all.
          >
          > That is not to say that pursuing ultimate resolution isn't worth while or
          > fascinating in and of itself but it is a very precise endeavor.
          >
          > NOTE:
          >
          > * Older objectives were designed for appropriately aged cedar oil with each
          > lens manufacturer providing their own optimum formulation. The index of
          > refraction of cedar oil increases with aging. Newer (does anyone know the
          > approximate transition date?) objectives were designed for Shillaber's (now
          > Cargille) non-drying synthetic oils, with a (sodium "D" line index of
          > refraction of 1.5150 at 25 deg C).
          >
          >
          > Randall
          >
          >
          >
          >
          >
          > -----Original Message-----
          > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
          > Behalf Of Charles Guevara
          > Sent: Tuesday, November 01, 2011 7:53 PM
          > To: Microscope@yahoogroups.com
          > Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
          > Plan-NeoFluar
          >
          >
          >    Thank you so much, Rich, for this practicle/hands on microscopy thread.
          >  
          >   Rolf, Randall, David have clarified performance for visual, vrs. image
          > capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
          > Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction collar
          > for coverslip/water colum variations?.
          >  
          >    Rolf noted that the specimen context: (wetmount-pondlife slides with
          > coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
          > noted that this context renders: 'Plan objectives utility as often over
          > rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
          > bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
          > though you do not like to muddle with oiled-optics...as a 'fall back...Rolf
          > suggested : water immersion/LOMO objectives.).
          >  
          >    I have : NA 1.25 condensors on both my work-horse stands....I have a
          > Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
          > objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
          > Nikon CFW 10X occulars with this setup.
          >  
          >  
          >     I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
          > with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars with
          > this setup.  
          >  
          >    Please...this is not a 'high-jack', of Rich's thread...please comment to
          > Rich (...err, to all)...although we rarely do it...why is not:
          > 'oil-bridgeing the substage-condensor not at least mentioned in this
          > discussion with Rich pondering objectives for his use( okay...Rich already
          > said he is not keen on imerrsion oil useage)?
          >  
          >    Please...as I have a variety of occulars...( okay...I think Rich already
          > said he is useing:'10X occulars'.)...please comment on how one need concern
          > a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
          > PlanApo?
          >  
          >  
          >     Do occulars even matter in Rich-guys setup?  With the very common: NA
          > 1.25 substage condensors...we often do not 'oil-bridge condensor to
          > slide'..does this factor in to Rich'es 'application Difference'?    thanks
          > for all this practicle/ applied microscopy, charlie guevara     (PS)  in
          > hobby astronomy...Apo refractors are costly, their proud owners seek out
          > planetary observing agendas for crisp/'tack-sharp'  color observations.  
          > Deep-sky (beyond our Suns family of interacting systems)...deep sky
          > observers always 'chant': so little color differences in 'deep sky
          > objects'...no need to worry about: 'purple fringes on bright stars...go
          > 'achro'...leave 'apos' for when you win the lottery.
          >
        • John
          ... Not exactly correct it doesn t have to be an apochromat to achieve 1.4na. It s a curious area no one makes apochromats any more only plan apochromats and
          Message 4 of 21 , Nov 3, 2011
            --- In Microscope@yahoogroups.com, "Selwyn St Leger" <selwynstleger@...> wrote:
            >
            > Thus, stopping down the condenser a little does not have a large effect on resolution. Also, apochromatic objectives (which they must be to attain 1.4 NA) because of their corrections require less stopping down of the condenser than achromats to achieve good contrast.
            >
            Not exactly correct it doesn't have to be an apochromat to achieve 1.4na. It's a curious area no one makes apochromats any more only plan apochromats and there are technical reasons why the colour focal plains of these are w shaped and also different for the 3 colours that they are measured with. This degrades resolution so in effect wont actually achieve 1.4na on axis when the best over all view is focused.

            I snipped the bit on shutting down the condenser aperture - but as it's nearly time for bed here. This increases contrast. Resolution really is contrast. There is plenty of information about mtf curves about on the web. These show the contrast of black and white lines as the spacing decreases. At 2na/lamda the contrast is 0. It's not a linear curve but at 2 twice that the contrast is 0.3 for a lens that meets raylieghs limit and nearly linear. It's 0.4 for an impossible perfect lens and less linear. As we don't usually look at black and white lines shutting down the condenser lens to boost contrast will inevitably show more detail. It's a pity some one can't post a picture of an objective working at it's max resolution. The na doesn't matter as all curves have the same properties. Defocus and the actual quality of the lens can change the results dramatically - both reduce contrast. Especially minimal amounts of defocus. Software can boost contrast and our eyes have curious properties in this area as well. basically it would be easy to cheat when taking a photo like this but it could get the point home.

            One aspect not mentioned is the slide thickness. Manufacturers oil's RI vary and I assume the required slide thicknesses do too. The thickness aspect is a bit like the cover slip thickness. The only one I have ever managed to tie down was Olympus, probably early and that stated 1mm. Defocusing the condenser also reduces it's na so the 1mm is probably rather critical as well.

            John
          • John
            Our eyes work the other way round - resolution goes down as contrast goes down but for some reason there is a odd kink at the bottom end that increases
            Message 5 of 21 , Nov 4, 2011
              Our eyes work the other way round - resolution goes down as contrast goes down but for some reason there is a odd kink at the bottom end that increases slightly at low contrast levels. Not that this matter much as the resolution at this point is about 0.1 lp/mm and the apparent contrast level is 0.1. When contrast is 1 we can manage 0.7 lp/mm. These are general figures that may well vary according to the individual. Where light levels should play a part is that they determine the diameter of the pupil.

              John

              --- In Microscope@yahoogroups.com, "rich_guy_looker" <rich_guy_looker@...> wrote:
              >
              >
              >
              > Resolution goes up
              > as contrast goes down, so;
              > theres' one good reason
              > why you should use a little light
              > as possible to light your specimen.
              >
              > It's all in the trade-offs
              > called "optical physics."
              >
              > As far as your comment
              > about slide thickness.....
              >
              > With tolerances of less than a few micrometers,
              > in both depth-of-field,
              > and proper alignment of the condenser output;
              > I'd bet you'd get a much better image and a
              > better-lighted specimen [especially for darkfield]
              > if you tried using precision-size cover-slips for
              > BOTH the slide that the specimen is placed on
              > AND the cover slip that's placed over
              > the specimen!
              >
              > Obviously, with these
              > small [micrometer] tolerances, the condenser output
              > would certainly be much less distorted
              > and better directed if it's only traveling
              > through a much thinner piece of glass that's
              > has a thickness accuracy of 0.005 millimeters!
              >
              > Rich
              >
            • Randall Buck
              Hi Selwyn, Thanks for your comments. Am I correct in paraphrasing, achieving optimum resolution is not the same as being able to use it in a meaningful way?
              Message 6 of 21 , Nov 4, 2011
                Hi Selwyn,

                Thanks for your comments. Am I correct in paraphrasing, achieving optimum
                resolution is not the same as being able to use it in a meaningful way?

                There is another aspect of this subject that I haven't "resolved". Using
                very high NA objectives (either dry or oil) with a wet-mount specimen does
                not seem to be practical due to what appears to be spherical aberration
                induced by the low index (~1.3) water layer under the coverslip. (
                Objectives designed for water immersion are the exception, of course. )

                On the other hand, diatom observers often use high index (higher than the
                1.52 of glass or immersion oil) mountant material (index ~2+) with great
                success.

                Do you have any thoughts on this matter?

                Best,

                Randall








                -----Original Message-----
                From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                Behalf Of Selwyn St Leger
                Sent: Thursday, November 03, 2011 11:23 AM
                To: Microscope@yahoogroups.com
                Subject: [Microscope] Re: Application Difference?? 100X PlanApo vs, 63X
                Plan-NeoFluar


                HiRandall,

                Those are interesting observations and well put.

                One point you raise merits further discussion. That is whether or not the
                condenser diaphragm should be reduced below NA 1.4 when employing a 1.4 NA
                objective.

                With geometric optics the condenser diaphragm is conjugate with (in focus
                at) the back focal plane of the objective. Thus, from this way of
                considering light transmission its effect is exactly the same as an
                equivalent stop in the back focal plane. However, as the image is formed
                from diffracted rays off the specimen some of these pass beyond the
                geometric light cone, impinge on the periphery of the objective to
                contribute to interference at the back focal plane.

                Thus, stopping down the condenser a little does not have a large effect on
                resolution. Also, apochromatic objectives (which they must be to attain 1.4
                NA) because of their corrections require less stopping down of the condenser
                than achromats to achieve good contrast.

                Nevertheless, your broader point has great merit. Attempting to get maximum
                resolution is not often, in contexts such as mine, worth the candle. Very
                high resolution has to be accompanied by the facility to interpret what has
                been resolved. That's fine with test diatoms but looking, say, at living
                cytoplasm in a protist there is not much point in being able to perceive a
                few more anonymous granules.

                Selwyn


                --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@...> wrote:
                >
                >
                > Hi,
                >
                > The more I think about the subject, the more I come to believe that John
                W.
                > (ajohn) has condensed the whole subject into a very succinct statement:
                > (Paraphrasing), Think of Very high NA objectives as though they are
                designed
                > for one thing only, viewing something that is attached to the underside of
                a
                > precision coverslip (one that meets the design specifications of the
                > objective in hand).
                >
                > You certainly don't want to be looking through anything other than
                immersion
                > oil * and the coverslip otherwise you will introduce an amount of
                spherical
                > aberration that will reduce the performance your expensive objective to
                that
                > of an objective for which you could have paid much less. Even mountant,
                > unless it is more of a filler than an intervening layer may require a
                > thinner coverslip to compensate for any added thickness.
                >
                > Also, remember that in order to take full advantage of the high NA of the
                > objective, you will need a high NA condenser. Since the condenser is
                > operating at high NA it requires improved optical correction to deliver a
                > precisely imaged beam to the subject plane. This is why the aplanatic
                > achromat was developed. Of course, it must be oiled to the bottom of the
                > slide, in fact, it will generally be impossible to align it of Kohler
                > without oiling. In addition, the iris should be used wide open, otherwise
                > your expensive 1.4NA aa condenser will be doing the job of a much less
                > expensive (lower NA) condenser and taking the performance of your High NA
                > objective down with it.
                >
                > This means that the perceived contrast improvement ordinarily gained by
                > reducing the condenser iris to 2/3 cannot be employed when the maximum
                > resolution of a high NA objective is the goal. In turn, this means that
                > the specimen itself must be of intrinsically high contrast (via staining,
                > etc) to be seen at all.
                >
                > That is not to say that pursuing ultimate resolution isn't worth while or
                > fascinating in and of itself but it is a very precise endeavor.
                >
                > NOTE:
                >
                > * Older objectives were designed for appropriately aged cedar oil with
                each
                > lens manufacturer providing their own optimum formulation. The index of
                > refraction of cedar oil increases with aging. Newer (does anyone know the
                > approximate transition date?) objectives were designed for Shillaber's
                (now
                > Cargille) non-drying synthetic oils, with a (sodium "D" line index of
                > refraction of 1.5150 at 25 deg C).
                >
                >
                > Randall
                >
                >
                >
                >
                >
                > -----Original Message-----
                > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                > Behalf Of Charles Guevara
                > Sent: Tuesday, November 01, 2011 7:53 PM
                > To: Microscope@yahoogroups.com
                > Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
                > Plan-NeoFluar
                >
                >
                >    Thank you so much, Rich, for this practicle/hands on microscopy thread.
                >  
                >   Rolf, Randall, David have clarified performance for visual, vrs. image
                > capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
                > Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction
                collar
                > for coverslip/water colum variations?.
                >  
                >    Rolf noted that the specimen context: (wetmount-pondlife slides with
                > coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
                > noted that this context renders: 'Plan objectives utility as often over
                > rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
                > bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
                > though you do not like to muddle with oiled-optics...as a 'fall
                back...Rolf
                > suggested : water immersion/LOMO objectives.).
                >  
                >    I have : NA 1.25 condensors on both my work-horse stands....I have a
                > Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
                > objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
                > Nikon CFW 10X occulars with this setup.
                >  
                >  
                >     I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
                > with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars
                with
                > this setup.  
                >  
                >    Please...this is not a 'high-jack', of Rich's thread...please comment
                to
                > Rich (...err, to all)...although we rarely do it...why is not:
                > 'oil-bridgeing the substage-condensor not at least mentioned in this
                > discussion with Rich pondering objectives for his use( okay...Rich already
                > said he is not keen on imerrsion oil useage)?
                >  
                >    Please...as I have a variety of occulars...( okay...I think Rich
                already
                > said he is useing:'10X occulars'.)...please comment on how one need
                concern
                > a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
                > PlanApo?
                >  
                >  
                >     Do occulars even matter in Rich-guys setup?  With the very common: NA
                > 1.25 substage condensors...we often do not 'oil-bridge condensor to
                > slide'..does this factor in to Rich'es 'application Difference'?    thanks
                > for all this practicle/ applied microscopy, charlie guevara     (PS)  in
                > hobby astronomy...Apo refractors are costly, their proud owners seek out
                > planetary observing agendas for crisp/'tack-sharp'  color observations.  
                > Deep-sky (beyond our Suns family of interacting systems)...deep sky
                > observers always 'chant': so little color differences in 'deep sky
                > objects'...no need to worry about: 'purple fringes on bright stars...go
                > 'achro'...leave 'apos' for when you win the lottery.
                >




                ------------------------------------

                Yahoo! Groups Links
              • John
                Me not Selwyn but I have. I m told that K K for instance uses thin cover slips to get round the high ri aspect. In some ways this relates to optical path
                Message 7 of 21 , Nov 4, 2011
                  Me not Selwyn but I have.

                  I'm told that K K for instance uses thin cover slips to get round the high ri aspect. In some ways this relates to optical path length which is a factor of length and RI. The detrimental effects are often very noticeable on strews that may (even often) have areas where there are 3 or 4 diatoms on top of each other plus mountant and cover slip. Even a normal 40x will not resolve the detail it should on many of them. This can even show up on K K's test slide in relationship to fully resolving pleurosigma angulatum as it's much thinner than some of the other diatoms and can finish up being too deep in the mountant to be easily and clearly resolved as it should be. Oddly an oil lens can help with this but in some ways that's logical. An oil lens expects a higher ri at it's end than a dry one. The effective OPL is there for more suitable for them. :-) That's my theory but so far it seems to be true but there is a fair chance NA has something to do with it as well. Basically higher contrast.

                  I've managed to get hold of some n0 0 slips but this still wont leave much room for excess mountant.

                  John

                  --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@...> wrote:
                  >
                  >
                  >
                  > Hi Selwyn,
                  >
                  > Thanks for your comments. Am I correct in paraphrasing, achieving optimum
                  > resolution is not the same as being able to use it in a meaningful way?
                  >
                  > There is another aspect of this subject that I haven't "resolved". Using
                  > very high NA objectives (either dry or oil) with a wet-mount specimen does
                  > not seem to be practical due to what appears to be spherical aberration
                  > induced by the low index (~1.3) water layer under the coverslip. (
                  > Objectives designed for water immersion are the exception, of course. )
                  >
                  > On the other hand, diatom observers often use high index (higher than the
                  > 1.52 of glass or immersion oil) mountant material (index ~2+) with great
                  > success.
                  >
                  > Do you have any thoughts on this matter?
                  >
                  > Best,
                  >
                  > Randall
                  >
                  >
                  >
                  >
                  >
                  >
                  >
                  >
                  > -----Original Message-----
                  > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                  > Behalf Of Selwyn St Leger
                  > Sent: Thursday, November 03, 2011 11:23 AM
                  > To: Microscope@yahoogroups.com
                  > Subject: [Microscope] Re: Application Difference?? 100X PlanApo vs, 63X
                  > Plan-NeoFluar
                  >
                  >
                  > HiRandall,
                  >
                  > Those are interesting observations and well put.
                  >
                  > One point you raise merits further discussion. That is whether or not the
                  > condenser diaphragm should be reduced below NA 1.4 when employing a 1.4 NA
                  > objective.
                  >
                  > With geometric optics the condenser diaphragm is conjugate with (in focus
                  > at) the back focal plane of the objective. Thus, from this way of
                  > considering light transmission its effect is exactly the same as an
                  > equivalent stop in the back focal plane. However, as the image is formed
                  > from diffracted rays off the specimen some of these pass beyond the
                  > geometric light cone, impinge on the periphery of the objective to
                  > contribute to interference at the back focal plane.
                  >
                  > Thus, stopping down the condenser a little does not have a large effect on
                  > resolution. Also, apochromatic objectives (which they must be to attain 1.4
                  > NA) because of their corrections require less stopping down of the condenser
                  > than achromats to achieve good contrast.
                  >
                  > Nevertheless, your broader point has great merit. Attempting to get maximum
                  > resolution is not often, in contexts such as mine, worth the candle. Very
                  > high resolution has to be accompanied by the facility to interpret what has
                  > been resolved. That's fine with test diatoms but looking, say, at living
                  > cytoplasm in a protist there is not much point in being able to perceive a
                  > few more anonymous granules.
                  >
                  > Selwyn
                  >
                  >
                  > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                  > >
                  > >
                  > > Hi,
                  > >
                  > > The more I think about the subject, the more I come to believe that John
                  > W.
                  > > (ajohn) has condensed the whole subject into a very succinct statement:
                  > > (Paraphrasing), Think of Very high NA objectives as though they are
                  > designed
                  > > for one thing only, viewing something that is attached to the underside of
                  > a
                  > > precision coverslip (one that meets the design specifications of the
                  > > objective in hand).
                  > >
                  > > You certainly don't want to be looking through anything other than
                  > immersion
                  > > oil * and the coverslip otherwise you will introduce an amount of
                  > spherical
                  > > aberration that will reduce the performance your expensive objective to
                  > that
                  > > of an objective for which you could have paid much less. Even mountant,
                  > > unless it is more of a filler than an intervening layer may require a
                  > > thinner coverslip to compensate for any added thickness.
                  > >
                  > > Also, remember that in order to take full advantage of the high NA of the
                  > > objective, you will need a high NA condenser. Since the condenser is
                  > > operating at high NA it requires improved optical correction to deliver a
                  > > precisely imaged beam to the subject plane. This is why the aplanatic
                  > > achromat was developed. Of course, it must be oiled to the bottom of the
                  > > slide, in fact, it will generally be impossible to align it of Kohler
                  > > without oiling. In addition, the iris should be used wide open, otherwise
                  > > your expensive 1.4NA aa condenser will be doing the job of a much less
                  > > expensive (lower NA) condenser and taking the performance of your High NA
                  > > objective down with it.
                  > >
                  > > This means that the perceived contrast improvement ordinarily gained by
                  > > reducing the condenser iris to 2/3 cannot be employed when the maximum
                  > > resolution of a high NA objective is the goal. In turn, this means that
                  > > the specimen itself must be of intrinsically high contrast (via staining,
                  > > etc) to be seen at all.
                  > >
                  > > That is not to say that pursuing ultimate resolution isn't worth while or
                  > > fascinating in and of itself but it is a very precise endeavor.
                  > >
                  > > NOTE:
                  > >
                  > > * Older objectives were designed for appropriately aged cedar oil with
                  > each
                  > > lens manufacturer providing their own optimum formulation. The index of
                  > > refraction of cedar oil increases with aging. Newer (does anyone know the
                  > > approximate transition date?) objectives were designed for Shillaber's
                  > (now
                  > > Cargille) non-drying synthetic oils, with a (sodium "D" line index of
                  > > refraction of 1.5150 at 25 deg C).
                  > >
                  > >
                  > > Randall
                  > >
                  > >
                  > >
                  > >
                  > >
                  > > -----Original Message-----
                  > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                  > > Behalf Of Charles Guevara
                  > > Sent: Tuesday, November 01, 2011 7:53 PM
                  > > To: Microscope@yahoogroups.com
                  > > Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
                  > > Plan-NeoFluar
                  > >
                  > >
                  > >    Thank you so much, Rich, for this practicle/hands on microscopy thread.
                  > >  
                  > >   Rolf, Randall, David have clarified performance for visual, vrs. image
                  > > capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
                  > > Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction
                  > collar
                  > > for coverslip/water colum variations?.
                  > >  
                  > >    Rolf noted that the specimen context: (wetmount-pondlife slides with
                  > > coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
                  > > noted that this context renders: 'Plan objectives utility as often over
                  > > rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
                  > > bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
                  > > though you do not like to muddle with oiled-optics...as a 'fall
                  > back...Rolf
                  > > suggested : water immersion/LOMO objectives.).
                  > >  
                  > >    I have : NA 1.25 condensors on both my work-horse stands....I have a
                  > > Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
                  > > objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
                  > > Nikon CFW 10X occulars with this setup.
                  > >  
                  > >  
                  > >     I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
                  > > with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars
                  > with
                  > > this setup.  
                  > >  
                  > >    Please...this is not a 'high-jack', of Rich's thread...please comment
                  > to
                  > > Rich (...err, to all)...although we rarely do it...why is not:
                  > > 'oil-bridgeing the substage-condensor not at least mentioned in this
                  > > discussion with Rich pondering objectives for his use( okay...Rich already
                  > > said he is not keen on imerrsion oil useage)?
                  > >  
                  > >    Please...as I have a variety of occulars...( okay...I think Rich
                  > already
                  > > said he is useing:'10X occulars'.)...please comment on how one need
                  > concern
                  > > a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
                  > > PlanApo?
                  > >  
                  > >  
                  > >     Do occulars even matter in Rich-guys setup?  With the very common: NA
                  > > 1.25 substage condensors...we often do not 'oil-bridge condensor to
                  > > slide'..does this factor in to Rich'es 'application Difference'?    thanks
                  > > for all this practicle/ applied microscopy, charlie guevara     (PS)  in
                  > > hobby astronomy...Apo refractors are costly, their proud owners seek out
                  > > planetary observing agendas for crisp/'tack-sharp'  color observations.  
                  > > Deep-sky (beyond our Suns family of interacting systems)...deep sky
                  > > observers always 'chant': so little color differences in 'deep sky
                  > > objects'...no need to worry about: 'purple fringes on bright stars...go
                  > > 'achro'...leave 'apos' for when you win the lottery.
                  > >
                  >
                  >
                  >
                  >
                  > ------------------------------------
                  >
                  > Yahoo! Groups Links
                  >
                • rene
                  I haven t followed the whole discussion, but my thoughts on the matter: Diatomists use thin coverglasses because it gives them most leeway. They generally go
                  Message 8 of 21 , Nov 4, 2011
                    I haven't followed the whole discussion, but my thoughts on the matter:

                    Diatomists use thin coverglasses because it gives them most leeway. They generally go to oil-immersion in the end, thin or thick coverslip
                    does not matter in practice. Most importantly, they put their material on the coverslip before mounting, the RI-influence of the mountant on spherical abberation is therefore minimal. Of course it would matter much oif you mount on the slide! Klaus Kemp makes test slides where he indeed put his specimen on the slide, and tries to make the mount as thin as possible. That's mainly because he wants the diatoms horizontally with the slide, and he cannot control that when he is going to mount a coverslip. IMHO I think that's wrong, but that's his choice. Besides, the test slide is a subjective test, not one valve is exactly the same as another. And he clearly states these slides should be used as a reference point: you know what it does with a certain lens so you can judge another lens.

                    As a last remark, a lot of people judge a car on the engine, regardless of the dirt road they live on.
                    What I mean is: there's a lot to be gained by good slide making practise.

                    FWIIW, René



                    --- In Microscope@yahoogroups.com, "John" <a.johnw@...> wrote:
                    >
                    > Me not Selwyn but I have.
                    >
                    > I'm told that K K for instance uses thin cover slips to get round the high ri aspect. In some ways this relates to optical path length which is a factor of length and RI. The detrimental effects are often very noticeable on strews that may (even often) have areas where there are 3 or 4 diatoms on top of each other plus mountant and cover slip. Even a normal 40x will not resolve the detail it should on many of them. This can even show up on K K's test slide in relationship to fully resolving pleurosigma angulatum as it's much thinner than some of the other diatoms and can finish up being too deep in the mountant to be easily and clearly resolved as it should be. Oddly an oil lens can help with this but in some ways that's logical. An oil lens expects a higher ri at it's end than a dry one. The effective OPL is there for more suitable for them. :-) That's my theory but so far it seems to be true but there is a fair chance NA has something to do with it as well. Basically higher contrast.
                    >
                    > I've managed to get hold of some n0 0 slips but this still wont leave much room for excess mountant.
                    >
                    > John
                    >
                    > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                    > >
                    > >
                    > >
                    > > Hi Selwyn,
                    > >
                    > > Thanks for your comments. Am I correct in paraphrasing, achieving optimum
                    > > resolution is not the same as being able to use it in a meaningful way?
                    > >
                    > > There is another aspect of this subject that I haven't "resolved". Using
                    > > very high NA objectives (either dry or oil) with a wet-mount specimen does
                    > > not seem to be practical due to what appears to be spherical aberration
                    > > induced by the low index (~1.3) water layer under the coverslip. (
                    > > Objectives designed for water immersion are the exception, of course. )
                    > >
                    > > On the other hand, diatom observers often use high index (higher than the
                    > > 1.52 of glass or immersion oil) mountant material (index ~2+) with great
                    > > success.
                    > >
                    > > Do you have any thoughts on this matter?
                    > >
                    > > Best,
                    > >
                    > > Randall
                    > >
                    > >
                    > >
                    > >
                    > >
                    > >
                    > >
                    > >
                    > > -----Original Message-----
                    > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                    > > Behalf Of Selwyn St Leger
                    > > Sent: Thursday, November 03, 2011 11:23 AM
                    > > To: Microscope@yahoogroups.com
                    > > Subject: [Microscope] Re: Application Difference?? 100X PlanApo vs, 63X
                    > > Plan-NeoFluar
                    > >
                    > >
                    > > HiRandall,
                    > >
                    > > Those are interesting observations and well put.
                    > >
                    > > One point you raise merits further discussion. That is whether or not the
                    > > condenser diaphragm should be reduced below NA 1.4 when employing a 1.4 NA
                    > > objective.
                    > >
                    > > With geometric optics the condenser diaphragm is conjugate with (in focus
                    > > at) the back focal plane of the objective. Thus, from this way of
                    > > considering light transmission its effect is exactly the same as an
                    > > equivalent stop in the back focal plane. However, as the image is formed
                    > > from diffracted rays off the specimen some of these pass beyond the
                    > > geometric light cone, impinge on the periphery of the objective to
                    > > contribute to interference at the back focal plane.
                    > >
                    > > Thus, stopping down the condenser a little does not have a large effect on
                    > > resolution. Also, apochromatic objectives (which they must be to attain 1.4
                    > > NA) because of their corrections require less stopping down of the condenser
                    > > than achromats to achieve good contrast.
                    > >
                    > > Nevertheless, your broader point has great merit. Attempting to get maximum
                    > > resolution is not often, in contexts such as mine, worth the candle. Very
                    > > high resolution has to be accompanied by the facility to interpret what has
                    > > been resolved. That's fine with test diatoms but looking, say, at living
                    > > cytoplasm in a protist there is not much point in being able to perceive a
                    > > few more anonymous granules.
                    > >
                    > > Selwyn
                    > >
                    > >
                    > > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                    > > >
                    > > >
                    > > > Hi,
                    > > >
                    > > > The more I think about the subject, the more I come to believe that John
                    > > W.
                    > > > (ajohn) has condensed the whole subject into a very succinct statement:
                    > > > (Paraphrasing), Think of Very high NA objectives as though they are
                    > > designed
                    > > > for one thing only, viewing something that is attached to the underside of
                    > > a
                    > > > precision coverslip (one that meets the design specifications of the
                    > > > objective in hand).
                    > > >
                    > > > You certainly don't want to be looking through anything other than
                    > > immersion
                    > > > oil * and the coverslip otherwise you will introduce an amount of
                    > > spherical
                    > > > aberration that will reduce the performance your expensive objective to
                    > > that
                    > > > of an objective for which you could have paid much less. Even mountant,
                    > > > unless it is more of a filler than an intervening layer may require a
                    > > > thinner coverslip to compensate for any added thickness.
                    > > >
                    > > > Also, remember that in order to take full advantage of the high NA of the
                    > > > objective, you will need a high NA condenser. Since the condenser is
                    > > > operating at high NA it requires improved optical correction to deliver a
                    > > > precisely imaged beam to the subject plane. This is why the aplanatic
                    > > > achromat was developed. Of course, it must be oiled to the bottom of the
                    > > > slide, in fact, it will generally be impossible to align it of Kohler
                    > > > without oiling. In addition, the iris should be used wide open, otherwise
                    > > > your expensive 1.4NA aa condenser will be doing the job of a much less
                    > > > expensive (lower NA) condenser and taking the performance of your High NA
                    > > > objective down with it.
                    > > >
                    > > > This means that the perceived contrast improvement ordinarily gained by
                    > > > reducing the condenser iris to 2/3 cannot be employed when the maximum
                    > > > resolution of a high NA objective is the goal. In turn, this means that
                    > > > the specimen itself must be of intrinsically high contrast (via staining,
                    > > > etc) to be seen at all.
                    > > >
                    > > > That is not to say that pursuing ultimate resolution isn't worth while or
                    > > > fascinating in and of itself but it is a very precise endeavor.
                    > > >
                    > > > NOTE:
                    > > >
                    > > > * Older objectives were designed for appropriately aged cedar oil with
                    > > each
                    > > > lens manufacturer providing their own optimum formulation. The index of
                    > > > refraction of cedar oil increases with aging. Newer (does anyone know the
                    > > > approximate transition date?) objectives were designed for Shillaber's
                    > > (now
                    > > > Cargille) non-drying synthetic oils, with a (sodium "D" line index of
                    > > > refraction of 1.5150 at 25 deg C).
                    > > >
                    > > >
                    > > > Randall
                    > > >
                    > > >
                    > > >
                    > > >
                    > > >
                    > > > -----Original Message-----
                    > > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                    > > > Behalf Of Charles Guevara
                    > > > Sent: Tuesday, November 01, 2011 7:53 PM
                    > > > To: Microscope@yahoogroups.com
                    > > > Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
                    > > > Plan-NeoFluar
                    > > >
                    > > >
                    > > >    Thank you so much, Rich, for this practicle/hands on microscopy thread.
                    > > >  
                    > > >   Rolf, Randall, David have clarified performance for visual, vrs. image
                    > > > capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
                    > > > Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction
                    > > collar
                    > > > for coverslip/water colum variations?.
                    > > >  
                    > > >    Rolf noted that the specimen context: (wetmount-pondlife slides with
                    > > > coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
                    > > > noted that this context renders: 'Plan objectives utility as often over
                    > > > rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
                    > > > bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
                    > > > though you do not like to muddle with oiled-optics...as a 'fall
                    > > back...Rolf
                    > > > suggested : water immersion/LOMO objectives.).
                    > > >  
                    > > >    I have : NA 1.25 condensors on both my work-horse stands....I have a
                    > > > Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
                    > > > objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
                    > > > Nikon CFW 10X occulars with this setup.
                    > > >  
                    > > >  
                    > > >     I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
                    > > > with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars
                    > > with
                    > > > this setup.  
                    > > >  
                    > > >    Please...this is not a 'high-jack', of Rich's thread...please comment
                    > > to
                    > > > Rich (...err, to all)...although we rarely do it...why is not:
                    > > > 'oil-bridgeing the substage-condensor not at least mentioned in this
                    > > > discussion with Rich pondering objectives for his use( okay...Rich already
                    > > > said he is not keen on imerrsion oil useage)?
                    > > >  
                    > > >    Please...as I have a variety of occulars...( okay...I think Rich
                    > > already
                    > > > said he is useing:'10X occulars'.)...please comment on how one need
                    > > concern
                    > > > a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
                    > > > PlanApo?
                    > > >  
                    > > >  
                    > > >     Do occulars even matter in Rich-guys setup?  With the very common: NA
                    > > > 1.25 substage condensors...we often do not 'oil-bridge condensor to
                    > > > slide'..does this factor in to Rich'es 'application Difference'?    thanks
                    > > > for all this practicle/ applied microscopy, charlie guevara     (PS)  in
                    > > > hobby astronomy...Apo refractors are costly, their proud owners seek out
                    > > > planetary observing agendas for crisp/'tack-sharp'  color observations.  
                    > > > Deep-sky (beyond our Suns family of interacting systems)...deep sky
                    > > > observers always 'chant': so little color differences in 'deep sky
                    > > > objects'...no need to worry about: 'purple fringes on bright stars...go
                    > > > 'achro'...leave 'apos' for when you win the lottery.
                    > > >
                    > >
                    > >
                    > >
                    > >
                    > > ------------------------------------
                    > >
                    > > Yahoo! Groups Links
                    > >
                    >
                  • John
                    I had a chat with Klaus Kemp last year Rene about mounting directly on a cover slip and he basically said no. I think the difficulties with mounting on the
                    Message 9 of 21 , Nov 4, 2011
                      I had a chat with Klaus Kemp last year Rene about mounting directly on a cover slip and he basically said no. I think the difficulties with mounting on the cover slip come about because there must be a layer of hi ri mountant between the slip and the diatom. This can be rather difficult to control. To much and things wont work out. I hope to continue the conversation at some point. He was about to give a lecture -:-) Not the best time to approach him. I missed an opportunity more recently. I'm told by some one who probably does know that he not only makes his preparations as thin as he can but also uses thin cover slips. I afraid I can't agree that this is just for the reason you mention so once again.

                      On my previous comment about opl basically neglecting dispersion if a 0.17mms cover slip was made of glass with and ri of 1.9 instead of about 1.55 it would behave optically as if it was 1.9/1.55 thicker about 0.2mms in this case. This will have an effect on a standard 40x objective if there is a significant amount in these terms of hi ri mountant under a cover slip. This is really of most importance when viewing what is considered to be a suitable test object for a 40x na 0.65 objective that will be near it's max capability in any case. It doesn't take much to mess things up. If you want to see the effect for your self buy a set of Brunel test strews. They don't cost much. Often rather than seeing nice straight striae all that will be seen is wavy lines or worse. I'm sure we have had this conversation before elsewhere. Curious fact though an oil lens will make a far better job of it. I don't think this is just down to higher na. It's also down to opl. Out of interest I could resolve all diatoms on KK's test slide but for some reason the PA I mentioned got more difficult. It's actual dots can be difficult anyway. Resolving just the grid pattern is a lot easier. I suspect Klauss does more than he lets on. He is a very capable diatom mounter.

                      As to Optical Path Length OPD might produce more meaningful google searches if interested. Optical Path Difference, a common way of optimising lenses. :-) If only it could be zero for all acceptance angles and colours.

                      One curious aspect of hi ri mountants is that there are claims that they increase the depth of field. Pass on that at the moment.

                      John

                      --- In Microscope@yahoogroups.com, "rene" <renevanwezel@...> wrote:
                      >
                      > I haven't followed the whole discussion, but my thoughts on the matter:
                      >
                      > Diatomists use thin coverglasses because it gives them most leeway. They generally go to oil-immersion in the end, thin or thick coverslip
                      > does not matter in practice. Most importantly, they put their material on the coverslip before mounting, the RI-influence of the mountant on spherical abberation is therefore minimal. Of course it would matter much oif you mount on the slide! Klaus Kemp makes test slides where he indeed put his specimen on the slide, and tries to make the mount as thin as possible. That's mainly because he wants the diatoms horizontally with the slide, and he cannot control that when he is going to mount a coverslip. IMHO I think that's wrong, but that's his choice. Besides, the test slide is a subjective test, not one valve is exactly the same as another. And he clearly states these slides should be used as a reference point: you know what it does with a certain lens so you can judge another lens.
                      >
                      > As a last remark, a lot of people judge a car on the engine, regardless of the dirt road they live on.
                      > What I mean is: there's a lot to be gained by good slide making practise.
                      >
                      > FWIIW, René
                      >
                      >
                      >
                      > --- In Microscope@yahoogroups.com, "John" <a.johnw@> wrote:
                      > >
                      > > Me not Selwyn but I have.
                      > >
                      > > I'm told that K K for instance uses thin cover slips to get round the high ri aspect. In some ways this relates to optical path length which is a factor of length and RI. The detrimental effects are often very noticeable on strews that may (even often) have areas where there are 3 or 4 diatoms on top of each other plus mountant and cover slip. Even a normal 40x will not resolve the detail it should on many of them. This can even show up on K K's test slide in relationship to fully resolving pleurosigma angulatum as it's much thinner than some of the other diatoms and can finish up being too deep in the mountant to be easily and clearly resolved as it should be. Oddly an oil lens can help with this but in some ways that's logical. An oil lens expects a higher ri at it's end than a dry one. The effective OPL is there for more suitable for them. :-) That's my theory but so far it seems to be true but there is a fair chance NA has something to do with it as well. Basically higher contrast.
                      > >
                      > > I've managed to get hold of some n0 0 slips but this still wont leave much room for excess mountant.
                      > >
                      > > John
                      > >
                      > > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                      > > >
                      > > >
                      > > >
                      > > > Hi Selwyn,
                      > > >
                      > > > Thanks for your comments. Am I correct in paraphrasing, achieving optimum
                      > > > resolution is not the same as being able to use it in a meaningful way?
                      > > >
                      > > > There is another aspect of this subject that I haven't "resolved". Using
                      > > > very high NA objectives (either dry or oil) with a wet-mount specimen does
                      > > > not seem to be practical due to what appears to be spherical aberration
                      > > > induced by the low index (~1.3) water layer under the coverslip. (
                      > > > Objectives designed for water immersion are the exception, of course. )
                      > > >
                      > > > On the other hand, diatom observers often use high index (higher than the
                      > > > 1.52 of glass or immersion oil) mountant material (index ~2+) with great
                      > > > success.
                      > > >
                      > > > Do you have any thoughts on this matter?
                      > > >
                      > > > Best,
                      > > >
                      > > > Randall
                      > > >
                      > > >
                      > > >
                      > > >
                      > > >
                      > > >
                      > > >
                      > > >
                      > > > -----Original Message-----
                      > > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                      > > > Behalf Of Selwyn St Leger
                      > > > Sent: Thursday, November 03, 2011 11:23 AM
                      > > > To: Microscope@yahoogroups.com
                      > > > Subject: [Microscope] Re: Application Difference?? 100X PlanApo vs, 63X
                      > > > Plan-NeoFluar
                      > > >
                      > > >
                      > > > HiRandall,
                      > > >
                      > > > Those are interesting observations and well put.
                      > > >
                      > > > One point you raise merits further discussion. That is whether or not the
                      > > > condenser diaphragm should be reduced below NA 1.4 when employing a 1.4 NA
                      > > > objective.
                      > > >
                      > > > With geometric optics the condenser diaphragm is conjugate with (in focus
                      > > > at) the back focal plane of the objective. Thus, from this way of
                      > > > considering light transmission its effect is exactly the same as an
                      > > > equivalent stop in the back focal plane. However, as the image is formed
                      > > > from diffracted rays off the specimen some of these pass beyond the
                      > > > geometric light cone, impinge on the periphery of the objective to
                      > > > contribute to interference at the back focal plane.
                      > > >
                      > > > Thus, stopping down the condenser a little does not have a large effect on
                      > > > resolution. Also, apochromatic objectives (which they must be to attain 1.4
                      > > > NA) because of their corrections require less stopping down of the condenser
                      > > > than achromats to achieve good contrast.
                      > > >
                      > > > Nevertheless, your broader point has great merit. Attempting to get maximum
                      > > > resolution is not often, in contexts such as mine, worth the candle. Very
                      > > > high resolution has to be accompanied by the facility to interpret what has
                      > > > been resolved. That's fine with test diatoms but looking, say, at living
                      > > > cytoplasm in a protist there is not much point in being able to perceive a
                      > > > few more anonymous granules.
                      > > >
                      > > > Selwyn
                      > > >
                      > > >
                      > > > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                      > > > >
                      > > > >
                      > > > > Hi,
                      > > > >
                      > > > > The more I think about the subject, the more I come to believe that John
                      > > > W.
                      > > > > (ajohn) has condensed the whole subject into a very succinct statement:
                      > > > > (Paraphrasing), Think of Very high NA objectives as though they are
                      > > > designed
                      > > > > for one thing only, viewing something that is attached to the underside of
                      > > > a
                      > > > > precision coverslip (one that meets the design specifications of the
                      > > > > objective in hand).
                      > > > >
                      > > > > You certainly don't want to be looking through anything other than
                      > > > immersion
                      > > > > oil * and the coverslip otherwise you will introduce an amount of
                      > > > spherical
                      > > > > aberration that will reduce the performance your expensive objective to
                      > > > that
                      > > > > of an objective for which you could have paid much less. Even mountant,
                      > > > > unless it is more of a filler than an intervening layer may require a
                      > > > > thinner coverslip to compensate for any added thickness.
                      > > > >
                      > > > > Also, remember that in order to take full advantage of the high NA of the
                      > > > > objective, you will need a high NA condenser. Since the condenser is
                      > > > > operating at high NA it requires improved optical correction to deliver a
                      > > > > precisely imaged beam to the subject plane. This is why the aplanatic
                      > > > > achromat was developed. Of course, it must be oiled to the bottom of the
                      > > > > slide, in fact, it will generally be impossible to align it of Kohler
                      > > > > without oiling. In addition, the iris should be used wide open, otherwise
                      > > > > your expensive 1.4NA aa condenser will be doing the job of a much less
                      > > > > expensive (lower NA) condenser and taking the performance of your High NA
                      > > > > objective down with it.
                      > > > >
                      > > > > This means that the perceived contrast improvement ordinarily gained by
                      > > > > reducing the condenser iris to 2/3 cannot be employed when the maximum
                      > > > > resolution of a high NA objective is the goal. In turn, this means that
                      > > > > the specimen itself must be of intrinsically high contrast (via staining,
                      > > > > etc) to be seen at all.
                      > > > >
                      > > > > That is not to say that pursuing ultimate resolution isn't worth while or
                      > > > > fascinating in and of itself but it is a very precise endeavor.
                      > > > >
                      > > > > NOTE:
                      > > > >
                      > > > > * Older objectives were designed for appropriately aged cedar oil with
                      > > > each
                      > > > > lens manufacturer providing their own optimum formulation. The index of
                      > > > > refraction of cedar oil increases with aging. Newer (does anyone know the
                      > > > > approximate transition date?) objectives were designed for Shillaber's
                      > > > (now
                      > > > > Cargille) non-drying synthetic oils, with a (sodium "D" line index of
                      > > > > refraction of 1.5150 at 25 deg C).
                      > > > >
                      > > > >
                      > > > > Randall
                      > > > >
                      > > > >
                      > > > >
                      > > > >
                      > > > >
                      > > > > -----Original Message-----
                      > > > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                      > > > > Behalf Of Charles Guevara
                      > > > > Sent: Tuesday, November 01, 2011 7:53 PM
                      > > > > To: Microscope@yahoogroups.com
                      > > > > Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
                      > > > > Plan-NeoFluar
                      > > > >
                      > > > >
                      > > > >    Thank you so much, Rich, for this practicle/hands on microscopy thread.
                      > > > >  
                      > > > >   Rolf, Randall, David have clarified performance for visual, vrs. image
                      > > > > capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
                      > > > > Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction
                      > > > collar
                      > > > > for coverslip/water colum variations?.
                      > > > >  
                      > > > >    Rolf noted that the specimen context: (wetmount-pondlife slides with
                      > > > > coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
                      > > > > noted that this context renders: 'Plan objectives utility as often over
                      > > > > rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
                      > > > > bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
                      > > > > though you do not like to muddle with oiled-optics...as a 'fall
                      > > > back...Rolf
                      > > > > suggested : water immersion/LOMO objectives.).
                      > > > >  
                      > > > >    I have : NA 1.25 condensors on both my work-horse stands....I have a
                      > > > > Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
                      > > > > objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
                      > > > > Nikon CFW 10X occulars with this setup.
                      > > > >  
                      > > > >  
                      > > > >     I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
                      > > > > with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars
                      > > > with
                      > > > > this setup.  
                      > > > >  
                      > > > >    Please...this is not a 'high-jack', of Rich's thread...please comment
                      > > > to
                      > > > > Rich (...err, to all)...although we rarely do it...why is not:
                      > > > > 'oil-bridgeing the substage-condensor not at least mentioned in this
                      > > > > discussion with Rich pondering objectives for his use( okay...Rich already
                      > > > > said he is not keen on imerrsion oil useage)?
                      > > > >  
                      > > > >    Please...as I have a variety of occulars...( okay...I think Rich
                      > > > already
                      > > > > said he is useing:'10X occulars'.)...please comment on how one need
                      > > > concern
                      > > > > a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
                      > > > > PlanApo?
                      > > > >  
                      > > > >  
                      > > > >     Do occulars even matter in Rich-guys setup?  With the very common: NA
                      > > > > 1.25 substage condensors...we often do not 'oil-bridge condensor to
                      > > > > slide'..does this factor in to Rich'es 'application Difference'?    thanks
                      > > > > for all this practicle/ applied microscopy, charlie guevara     (PS)  in
                      > > > > hobby astronomy...Apo refractors are costly, their proud owners seek out
                      > > > > planetary observing agendas for crisp/'tack-sharp'  color observations.  
                      > > > > Deep-sky (beyond our Suns family of interacting systems)...deep sky
                      > > > > observers always 'chant': so little color differences in 'deep sky
                      > > > > objects'...no need to worry about: 'purple fringes on bright stars...go
                      > > > > 'achro'...leave 'apos' for when you win the lottery.
                      > > > >
                      > > >
                      > > >
                      > > >
                      > > >
                      > > > ------------------------------------
                      > > >
                      > > > Yahoo! Groups Links
                      > > >
                      > >
                      >
                    • rene
                      hmm, this was also in discussion with him, but maybe I just thought he agreed with my thinking ;-) Well, I ll see him again next week, will ask him again about
                      Message 10 of 21 , Nov 4, 2011
                        hmm, this was also in discussion with him, but maybe I just thought he agreed with my thinking ;-) Well, I'll see him again next week, will ask him again about it. It might as well have something to do with the orientation of the valve, and that it is easier to lay it up or down on the slide. To be honest, I have no idea what way the valve should be 'up'.

                        Concerning the DOF, like you say a .17 coverslip with RI 1.9 would be optically .04mm thicker then one of RI 1.51. This effect also increases the DOF. However, I can't say I have seen that for myself, as the DOF of high NA objective is already very close to zero in any case. Would make for a nice experiment, anyone?

                        Best wishes, René


                        --- In Microscope@yahoogroups.com, "John" <a.johnw@...> wrote:
                        >
                        > I had a chat with Klaus Kemp last year Rene about mounting directly on a cover slip and he basically said no. I think the difficulties with mounting on the cover slip come about because there must be a layer of hi ri mountant between the slip and the diatom. This can be rather difficult to control. To much and things wont work out. I hope to continue the conversation at some point. He was about to give a lecture -:-) Not the best time to approach him. I missed an opportunity more recently. I'm told by some one who probably does know that he not only makes his preparations as thin as he can but also uses thin cover slips. I afraid I can't agree that this is just for the reason you mention so once again.
                        >
                        > On my previous comment about opl basically neglecting dispersion if a 0.17mms cover slip was made of glass with and ri of 1.9 instead of about 1.55 it would behave optically as if it was 1.9/1.55 thicker about 0.2mms in this case. This will have an effect on a standard 40x objective if there is a significant amount in these terms of hi ri mountant under a cover slip. This is really of most importance when viewing what is considered to be a suitable test object for a 40x na 0.65 objective that will be near it's max capability in any case. It doesn't take much to mess things up. If you want to see the effect for your self buy a set of Brunel test strews. They don't cost much. Often rather than seeing nice straight striae all that will be seen is wavy lines or worse. I'm sure we have had this conversation before elsewhere. Curious fact though an oil lens will make a far better job of it. I don't think this is just down to higher na. It's also down to opl. Out of interest I could resolve all diatoms on KK's test slide but for some reason the PA I mentioned got more difficult. It's actual dots can be difficult anyway. Resolving just the grid pattern is a lot easier. I suspect Klauss does more than he lets on. He is a very capable diatom mounter.
                        >
                        > As to Optical Path Length OPD might produce more meaningful google searches if interested. Optical Path Difference, a common way of optimising lenses. :-) If only it could be zero for all acceptance angles and colours.
                        >
                        > One curious aspect of hi ri mountants is that there are claims that they increase the depth of field. Pass on that at the moment.
                        >
                        > John
                        >
                        > --- In Microscope@yahoogroups.com, "rene" <renevanwezel@> wrote:
                        > >
                        > > I haven't followed the whole discussion, but my thoughts on the matter:
                        > >
                        > > Diatomists use thin coverglasses because it gives them most leeway. They generally go to oil-immersion in the end, thin or thick coverslip
                        > > does not matter in practice. Most importantly, they put their material on the coverslip before mounting, the RI-influence of the mountant on spherical abberation is therefore minimal. Of course it would matter much oif you mount on the slide! Klaus Kemp makes test slides where he indeed put his specimen on the slide, and tries to make the mount as thin as possible. That's mainly because he wants the diatoms horizontally with the slide, and he cannot control that when he is going to mount a coverslip. IMHO I think that's wrong, but that's his choice. Besides, the test slide is a subjective test, not one valve is exactly the same as another. And he clearly states these slides should be used as a reference point: you know what it does with a certain lens so you can judge another lens.
                        > >
                        > > As a last remark, a lot of people judge a car on the engine, regardless of the dirt road they live on.
                        > > What I mean is: there's a lot to be gained by good slide making practise.
                        > >
                        > > FWIIW, René
                        > >
                        > >
                        > >
                        > > --- In Microscope@yahoogroups.com, "John" <a.johnw@> wrote:
                        > > >
                        > > > Me not Selwyn but I have.
                        > > >
                        > > > I'm told that K K for instance uses thin cover slips to get round the high ri aspect. In some ways this relates to optical path length which is a factor of length and RI. The detrimental effects are often very noticeable on strews that may (even often) have areas where there are 3 or 4 diatoms on top of each other plus mountant and cover slip. Even a normal 40x will not resolve the detail it should on many of them. This can even show up on K K's test slide in relationship to fully resolving pleurosigma angulatum as it's much thinner than some of the other diatoms and can finish up being too deep in the mountant to be easily and clearly resolved as it should be. Oddly an oil lens can help with this but in some ways that's logical. An oil lens expects a higher ri at it's end than a dry one. The effective OPL is there for more suitable for them. :-) That's my theory but so far it seems to be true but there is a fair chance NA has something to do with it as well. Basically higher contrast.
                        > > >
                        > > > I've managed to get hold of some n0 0 slips but this still wont leave much room for excess mountant.
                        > > >
                        > > > John
                        > > >
                        > > > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                        > > > >
                        > > > >
                        > > > >
                        > > > > Hi Selwyn,
                        > > > >
                        > > > > Thanks for your comments. Am I correct in paraphrasing, achieving optimum
                        > > > > resolution is not the same as being able to use it in a meaningful way?
                        > > > >
                        > > > > There is another aspect of this subject that I haven't "resolved". Using
                        > > > > very high NA objectives (either dry or oil) with a wet-mount specimen does
                        > > > > not seem to be practical due to what appears to be spherical aberration
                        > > > > induced by the low index (~1.3) water layer under the coverslip. (
                        > > > > Objectives designed for water immersion are the exception, of course. )
                        > > > >
                        > > > > On the other hand, diatom observers often use high index (higher than the
                        > > > > 1.52 of glass or immersion oil) mountant material (index ~2+) with great
                        > > > > success.
                        > > > >
                        > > > > Do you have any thoughts on this matter?
                        > > > >
                        > > > > Best,
                        > > > >
                        > > > > Randall
                        > > > >
                        > > > >
                        > > > >
                        > > > >
                        > > > >
                        > > > >
                        > > > >
                        > > > >
                        > > > > -----Original Message-----
                        > > > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                        > > > > Behalf Of Selwyn St Leger
                        > > > > Sent: Thursday, November 03, 2011 11:23 AM
                        > > > > To: Microscope@yahoogroups.com
                        > > > > Subject: [Microscope] Re: Application Difference?? 100X PlanApo vs, 63X
                        > > > > Plan-NeoFluar
                        > > > >
                        > > > >
                        > > > > HiRandall,
                        > > > >
                        > > > > Those are interesting observations and well put.
                        > > > >
                        > > > > One point you raise merits further discussion. That is whether or not the
                        > > > > condenser diaphragm should be reduced below NA 1.4 when employing a 1.4 NA
                        > > > > objective.
                        > > > >
                        > > > > With geometric optics the condenser diaphragm is conjugate with (in focus
                        > > > > at) the back focal plane of the objective. Thus, from this way of
                        > > > > considering light transmission its effect is exactly the same as an
                        > > > > equivalent stop in the back focal plane. However, as the image is formed
                        > > > > from diffracted rays off the specimen some of these pass beyond the
                        > > > > geometric light cone, impinge on the periphery of the objective to
                        > > > > contribute to interference at the back focal plane.
                        > > > >
                        > > > > Thus, stopping down the condenser a little does not have a large effect on
                        > > > > resolution. Also, apochromatic objectives (which they must be to attain 1.4
                        > > > > NA) because of their corrections require less stopping down of the condenser
                        > > > > than achromats to achieve good contrast.
                        > > > >
                        > > > > Nevertheless, your broader point has great merit. Attempting to get maximum
                        > > > > resolution is not often, in contexts such as mine, worth the candle. Very
                        > > > > high resolution has to be accompanied by the facility to interpret what has
                        > > > > been resolved. That's fine with test diatoms but looking, say, at living
                        > > > > cytoplasm in a protist there is not much point in being able to perceive a
                        > > > > few more anonymous granules.
                        > > > >
                        > > > > Selwyn
                        > > > >
                        > > > >
                        > > > > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                        > > > > >
                        > > > > >
                        > > > > > Hi,
                        > > > > >
                        > > > > > The more I think about the subject, the more I come to believe that John
                        > > > > W.
                        > > > > > (ajohn) has condensed the whole subject into a very succinct statement:
                        > > > > > (Paraphrasing), Think of Very high NA objectives as though they are
                        > > > > designed
                        > > > > > for one thing only, viewing something that is attached to the underside of
                        > > > > a
                        > > > > > precision coverslip (one that meets the design specifications of the
                        > > > > > objective in hand).
                        > > > > >
                        > > > > > You certainly don't want to be looking through anything other than
                        > > > > immersion
                        > > > > > oil * and the coverslip otherwise you will introduce an amount of
                        > > > > spherical
                        > > > > > aberration that will reduce the performance your expensive objective to
                        > > > > that
                        > > > > > of an objective for which you could have paid much less. Even mountant,
                        > > > > > unless it is more of a filler than an intervening layer may require a
                        > > > > > thinner coverslip to compensate for any added thickness.
                        > > > > >
                        > > > > > Also, remember that in order to take full advantage of the high NA of the
                        > > > > > objective, you will need a high NA condenser. Since the condenser is
                        > > > > > operating at high NA it requires improved optical correction to deliver a
                        > > > > > precisely imaged beam to the subject plane. This is why the aplanatic
                        > > > > > achromat was developed. Of course, it must be oiled to the bottom of the
                        > > > > > slide, in fact, it will generally be impossible to align it of Kohler
                        > > > > > without oiling. In addition, the iris should be used wide open, otherwise
                        > > > > > your expensive 1.4NA aa condenser will be doing the job of a much less
                        > > > > > expensive (lower NA) condenser and taking the performance of your High NA
                        > > > > > objective down with it.
                        > > > > >
                        > > > > > This means that the perceived contrast improvement ordinarily gained by
                        > > > > > reducing the condenser iris to 2/3 cannot be employed when the maximum
                        > > > > > resolution of a high NA objective is the goal. In turn, this means that
                        > > > > > the specimen itself must be of intrinsically high contrast (via staining,
                        > > > > > etc) to be seen at all.
                        > > > > >
                        > > > > > That is not to say that pursuing ultimate resolution isn't worth while or
                        > > > > > fascinating in and of itself but it is a very precise endeavor.
                        > > > > >
                        > > > > > NOTE:
                        > > > > >
                        > > > > > * Older objectives were designed for appropriately aged cedar oil with
                        > > > > each
                        > > > > > lens manufacturer providing their own optimum formulation. The index of
                        > > > > > refraction of cedar oil increases with aging. Newer (does anyone know the
                        > > > > > approximate transition date?) objectives were designed for Shillaber's
                        > > > > (now
                        > > > > > Cargille) non-drying synthetic oils, with a (sodium "D" line index of
                        > > > > > refraction of 1.5150 at 25 deg C).
                        > > > > >
                        > > > > >
                        > > > > > Randall
                        > > > > >
                        > > > > >
                        > > > > >
                        > > > > >
                        > > > > >
                        > > > > > -----Original Message-----
                        > > > > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                        > > > > > Behalf Of Charles Guevara
                        > > > > > Sent: Tuesday, November 01, 2011 7:53 PM
                        > > > > > To: Microscope@yahoogroups.com
                        > > > > > Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
                        > > > > > Plan-NeoFluar
                        > > > > >
                        > > > > >
                        > > > > >    Thank you so much, Rich, for this practicle/hands on microscopy thread.
                        > > > > >  
                        > > > > >   Rolf, Randall, David have clarified performance for visual, vrs. image
                        > > > > > capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
                        > > > > > Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction
                        > > > > collar
                        > > > > > for coverslip/water colum variations?.
                        > > > > >  
                        > > > > >    Rolf noted that the specimen context: (wetmount-pondlife slides with
                        > > > > > coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
                        > > > > > noted that this context renders: 'Plan objectives utility as often over
                        > > > > > rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
                        > > > > > bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
                        > > > > > though you do not like to muddle with oiled-optics...as a 'fall
                        > > > > back...Rolf
                        > > > > > suggested : water immersion/LOMO objectives.).
                        > > > > >  
                        > > > > >    I have : NA 1.25 condensors on both my work-horse stands....I have a
                        > > > > > Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
                        > > > > > objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
                        > > > > > Nikon CFW 10X occulars with this setup.
                        > > > > >  
                        > > > > >  
                        > > > > >     I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
                        > > > > > with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars
                        > > > > with
                        > > > > > this setup.  
                        > > > > >  
                        > > > > >    Please...this is not a 'high-jack', of Rich's thread...please comment
                        > > > > to
                        > > > > > Rich (...err, to all)...although we rarely do it...why is not:
                        > > > > > 'oil-bridgeing the substage-condensor not at least mentioned in this
                        > > > > > discussion with Rich pondering objectives for his use( okay...Rich already
                        > > > > > said he is not keen on imerrsion oil useage)?
                        > > > > >  
                        > > > > >    Please...as I have a variety of occulars...( okay...I think Rich
                        > > > > already
                        > > > > > said he is useing:'10X occulars'.)...please comment on how one need
                        > > > > concern
                        > > > > > a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
                        > > > > > PlanApo?
                        > > > > >  
                        > > > > >  
                        > > > > >     Do occulars even matter in Rich-guys setup?  With the very common: NA
                        > > > > > 1.25 substage condensors...we often do not 'oil-bridge condensor to
                        > > > > > slide'..does this factor in to Rich'es 'application Difference'?    thanks
                        > > > > > for all this practicle/ applied microscopy, charlie guevara     (PS)  in
                        > > > > > hobby astronomy...Apo refractors are costly, their proud owners seek out
                        > > > > > planetary observing agendas for crisp/'tack-sharp'  color observations.  
                        > > > > > Deep-sky (beyond our Suns family of interacting systems)...deep sky
                        > > > > > observers always 'chant': so little color differences in 'deep sky
                        > > > > > objects'...no need to worry about: 'purple fringes on bright stars...go
                        > > > > > 'achro'...leave 'apos' for when you win the lottery.
                        > > > > >
                        > > > >
                        > > > >
                        > > > >
                        > > > >
                        > > > > ------------------------------------
                        > > > >
                        > > > > Yahoo! Groups Links
                        > > > >
                        > > >
                        > >
                        >
                      • John
                        I don t get the dof either Rene and can t say I have noticed anything. I would expect the rays to bend more sharply as the na is higher but sometimes things
                        Message 11 of 21 , Nov 5, 2011
                          I don't get the dof either Rene and can't say I have noticed anything. I would expect the rays to bend more sharply as the na is higher but sometimes things like this aren't intuitive. I do have some A P in balsam. :-) Not the ideal subject for comparing. Can't even find them in transmitted light, visible if that's the correct word with crossed polarisers and a lot more so with dic.

                          On up and down sides I have heard that when Klauss does a demo he can turn them over. If you manage to clarify that aspect maybe you could let me know. I only manage see him once a year. He usually attends one of the microscope meets in the uk. I have some A P to mount so may find out myself at some point but at the moment other things are more pressing. I also have samples of several species I have collected myself - I need to re digest them though as some algae has formed. I had reasons for not wanting to keep them in alcohol and should have added a drop of formalin.

                          John

                          --- In Microscope@yahoogroups.com, "rene" <renevanwezel@...> wrote:
                          >
                          > hmm, this was also in discussion with him, but maybe I just thought he agreed with my thinking ;-) Well, I'll see him again next week, will ask him again about it. It might as well have something to do with the orientation of the valve, and that it is easier to lay it up or down on the slide. To be honest, I have no idea what way the valve should be 'up'.
                          >
                          > Concerning the DOF, like you say a .17 coverslip with RI 1.9 would be optically .04mm thicker then one of RI 1.51. This effect also increases the DOF. However, I can't say I have seen that for myself, as the DOF of high NA objective is already very close to zero in any case. Would make for a nice experiment, anyone?
                          >
                          > Best wishes, René
                          >
                          >
                          > --- In Microscope@yahoogroups.com, "John" <a.johnw@> wrote:
                          > >
                          > > I had a chat with Klaus Kemp last year Rene about mounting directly on a cover slip and he basically said no. I think the difficulties with mounting on the cover slip come about because there must be a layer of hi ri mountant between the slip and the diatom. This can be rather difficult to control. To much and things wont work out. I hope to continue the conversation at some point. He was about to give a lecture -:-) Not the best time to approach him. I missed an opportunity more recently. I'm told by some one who probably does know that he not only makes his preparations as thin as he can but also uses thin cover slips. I afraid I can't agree that this is just for the reason you mention so once again.
                          > >
                          > > On my previous comment about opl basically neglecting dispersion if a 0.17mms cover slip was made of glass with and ri of 1.9 instead of about 1.55 it would behave optically as if it was 1.9/1.55 thicker about 0.2mms in this case. This will have an effect on a standard 40x objective if there is a significant amount in these terms of hi ri mountant under a cover slip. This is really of most importance when viewing what is considered to be a suitable test object for a 40x na 0.65 objective that will be near it's max capability in any case. It doesn't take much to mess things up. If you want to see the effect for your self buy a set of Brunel test strews. They don't cost much. Often rather than seeing nice straight striae all that will be seen is wavy lines or worse. I'm sure we have had this conversation before elsewhere. Curious fact though an oil lens will make a far better job of it. I don't think this is just down to higher na. It's also down to opl. Out of interest I could resolve all diatoms on KK's test slide but for some reason the PA I mentioned got more difficult. It's actual dots can be difficult anyway. Resolving just the grid pattern is a lot easier. I suspect Klauss does more than he lets on. He is a very capable diatom mounter.
                          > >
                          > > As to Optical Path Length OPD might produce more meaningful google searches if interested. Optical Path Difference, a common way of optimising lenses. :-) If only it could be zero for all acceptance angles and colours.
                          > >
                          > > One curious aspect of hi ri mountants is that there are claims that they increase the depth of field. Pass on that at the moment.
                          > >
                          > > John
                          > >
                          > > --- In Microscope@yahoogroups.com, "rene" <renevanwezel@> wrote:
                          > > >
                          > > > I haven't followed the whole discussion, but my thoughts on the matter:
                          > > >
                          > > > Diatomists use thin coverglasses because it gives them most leeway. They generally go to oil-immersion in the end, thin or thick coverslip
                          > > > does not matter in practice. Most importantly, they put their material on the coverslip before mounting, the RI-influence of the mountant on spherical abberation is therefore minimal. Of course it would matter much oif you mount on the slide! Klaus Kemp makes test slides where he indeed put his specimen on the slide, and tries to make the mount as thin as possible. That's mainly because he wants the diatoms horizontally with the slide, and he cannot control that when he is going to mount a coverslip. IMHO I think that's wrong, but that's his choice. Besides, the test slide is a subjective test, not one valve is exactly the same as another. And he clearly states these slides should be used as a reference point: you know what it does with a certain lens so you can judge another lens.
                          > > >
                          > > > As a last remark, a lot of people judge a car on the engine, regardless of the dirt road they live on.
                          > > > What I mean is: there's a lot to be gained by good slide making practise.
                          > > >
                          > > > FWIIW, René
                          > > >
                          > > >
                          > > >
                          > > > --- In Microscope@yahoogroups.com, "John" <a.johnw@> wrote:
                          > > > >
                          > > > > Me not Selwyn but I have.
                          > > > >
                          > > > > I'm told that K K for instance uses thin cover slips to get round the high ri aspect. In some ways this relates to optical path length which is a factor of length and RI. The detrimental effects are often very noticeable on strews that may (even often) have areas where there are 3 or 4 diatoms on top of each other plus mountant and cover slip. Even a normal 40x will not resolve the detail it should on many of them. This can even show up on K K's test slide in relationship to fully resolving pleurosigma angulatum as it's much thinner than some of the other diatoms and can finish up being too deep in the mountant to be easily and clearly resolved as it should be. Oddly an oil lens can help with this but in some ways that's logical. An oil lens expects a higher ri at it's end than a dry one. The effective OPL is there for more suitable for them. :-) That's my theory but so far it seems to be true but there is a fair chance NA has something to do with it as well. Basically higher contrast.
                          > > > >
                          > > > > I've managed to get hold of some n0 0 slips but this still wont leave much room for excess mountant.
                          > > > >
                          > > > > John
                          > > > >
                          > > > > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > > Hi Selwyn,
                          > > > > >
                          > > > > > Thanks for your comments. Am I correct in paraphrasing, achieving optimum
                          > > > > > resolution is not the same as being able to use it in a meaningful way?
                          > > > > >
                          > > > > > There is another aspect of this subject that I haven't "resolved". Using
                          > > > > > very high NA objectives (either dry or oil) with a wet-mount specimen does
                          > > > > > not seem to be practical due to what appears to be spherical aberration
                          > > > > > induced by the low index (~1.3) water layer under the coverslip. (
                          > > > > > Objectives designed for water immersion are the exception, of course. )
                          > > > > >
                          > > > > > On the other hand, diatom observers often use high index (higher than the
                          > > > > > 1.52 of glass or immersion oil) mountant material (index ~2+) with great
                          > > > > > success.
                          > > > > >
                          > > > > > Do you have any thoughts on this matter?
                          > > > > >
                          > > > > > Best,
                          > > > > >
                          > > > > > Randall
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > > -----Original Message-----
                          > > > > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                          > > > > > Behalf Of Selwyn St Leger
                          > > > > > Sent: Thursday, November 03, 2011 11:23 AM
                          > > > > > To: Microscope@yahoogroups.com
                          > > > > > Subject: [Microscope] Re: Application Difference?? 100X PlanApo vs, 63X
                          > > > > > Plan-NeoFluar
                          > > > > >
                          > > > > >
                          > > > > > HiRandall,
                          > > > > >
                          > > > > > Those are interesting observations and well put.
                          > > > > >
                          > > > > > One point you raise merits further discussion. That is whether or not the
                          > > > > > condenser diaphragm should be reduced below NA 1.4 when employing a 1.4 NA
                          > > > > > objective.
                          > > > > >
                          > > > > > With geometric optics the condenser diaphragm is conjugate with (in focus
                          > > > > > at) the back focal plane of the objective. Thus, from this way of
                          > > > > > considering light transmission its effect is exactly the same as an
                          > > > > > equivalent stop in the back focal plane. However, as the image is formed
                          > > > > > from diffracted rays off the specimen some of these pass beyond the
                          > > > > > geometric light cone, impinge on the periphery of the objective to
                          > > > > > contribute to interference at the back focal plane.
                          > > > > >
                          > > > > > Thus, stopping down the condenser a little does not have a large effect on
                          > > > > > resolution. Also, apochromatic objectives (which they must be to attain 1.4
                          > > > > > NA) because of their corrections require less stopping down of the condenser
                          > > > > > than achromats to achieve good contrast.
                          > > > > >
                          > > > > > Nevertheless, your broader point has great merit. Attempting to get maximum
                          > > > > > resolution is not often, in contexts such as mine, worth the candle. Very
                          > > > > > high resolution has to be accompanied by the facility to interpret what has
                          > > > > > been resolved. That's fine with test diatoms but looking, say, at living
                          > > > > > cytoplasm in a protist there is not much point in being able to perceive a
                          > > > > > few more anonymous granules.
                          > > > > >
                          > > > > > Selwyn
                          > > > > >
                          > > > > >
                          > > > > > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                          > > > > > >
                          > > > > > >
                          > > > > > > Hi,
                          > > > > > >
                          > > > > > > The more I think about the subject, the more I come to believe that John
                          > > > > > W.
                          > > > > > > (ajohn) has condensed the whole subject into a very succinct statement:
                          > > > > > > (Paraphrasing), Think of Very high NA objectives as though they are
                          > > > > > designed
                          > > > > > > for one thing only, viewing something that is attached to the underside of
                          > > > > > a
                          > > > > > > precision coverslip (one that meets the design specifications of the
                          > > > > > > objective in hand).
                          > > > > > >
                          > > > > > > You certainly don't want to be looking through anything other than
                          > > > > > immersion
                          > > > > > > oil * and the coverslip otherwise you will introduce an amount of
                          > > > > > spherical
                          > > > > > > aberration that will reduce the performance your expensive objective to
                          > > > > > that
                          > > > > > > of an objective for which you could have paid much less. Even mountant,
                          > > > > > > unless it is more of a filler than an intervening layer may require a
                          > > > > > > thinner coverslip to compensate for any added thickness.
                          > > > > > >
                          > > > > > > Also, remember that in order to take full advantage of the high NA of the
                          > > > > > > objective, you will need a high NA condenser. Since the condenser is
                          > > > > > > operating at high NA it requires improved optical correction to deliver a
                          > > > > > > precisely imaged beam to the subject plane. This is why the aplanatic
                          > > > > > > achromat was developed. Of course, it must be oiled to the bottom of the
                          > > > > > > slide, in fact, it will generally be impossible to align it of Kohler
                          > > > > > > without oiling. In addition, the iris should be used wide open, otherwise
                          > > > > > > your expensive 1.4NA aa condenser will be doing the job of a much less
                          > > > > > > expensive (lower NA) condenser and taking the performance of your High NA
                          > > > > > > objective down with it.
                          > > > > > >
                          > > > > > > This means that the perceived contrast improvement ordinarily gained by
                          > > > > > > reducing the condenser iris to 2/3 cannot be employed when the maximum
                          > > > > > > resolution of a high NA objective is the goal. In turn, this means that
                          > > > > > > the specimen itself must be of intrinsically high contrast (via staining,
                          > > > > > > etc) to be seen at all.
                          > > > > > >
                          > > > > > > That is not to say that pursuing ultimate resolution isn't worth while or
                          > > > > > > fascinating in and of itself but it is a very precise endeavor.
                          > > > > > >
                          > > > > > > NOTE:
                          > > > > > >
                          > > > > > > * Older objectives were designed for appropriately aged cedar oil with
                          > > > > > each
                          > > > > > > lens manufacturer providing their own optimum formulation. The index of
                          > > > > > > refraction of cedar oil increases with aging. Newer (does anyone know the
                          > > > > > > approximate transition date?) objectives were designed for Shillaber's
                          > > > > > (now
                          > > > > > > Cargille) non-drying synthetic oils, with a (sodium "D" line index of
                          > > > > > > refraction of 1.5150 at 25 deg C).
                          > > > > > >
                          > > > > > >
                          > > > > > > Randall
                          > > > > > >
                          > > > > > >
                          > > > > > >
                          > > > > > >
                          > > > > > >
                          > > > > > > -----Original Message-----
                          > > > > > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                          > > > > > > Behalf Of Charles Guevara
                          > > > > > > Sent: Tuesday, November 01, 2011 7:53 PM
                          > > > > > > To: Microscope@yahoogroups.com
                          > > > > > > Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
                          > > > > > > Plan-NeoFluar
                          > > > > > >
                          > > > > > >
                          > > > > > >    Thank you so much, Rich, for this practicle/hands on microscopy thread.
                          > > > > > >  
                          > > > > > >   Rolf, Randall, David have clarified performance for visual, vrs. image
                          > > > > > > capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
                          > > > > > > Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction
                          > > > > > collar
                          > > > > > > for coverslip/water colum variations?.
                          > > > > > >  
                          > > > > > >    Rolf noted that the specimen context: (wetmount-pondlife slides with
                          > > > > > > coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
                          > > > > > > noted that this context renders: 'Plan objectives utility as often over
                          > > > > > > rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
                          > > > > > > bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
                          > > > > > > though you do not like to muddle with oiled-optics...as a 'fall
                          > > > > > back...Rolf
                          > > > > > > suggested : water immersion/LOMO objectives.).
                          > > > > > >  
                          > > > > > >    I have : NA 1.25 condensors on both my work-horse stands....I have a
                          > > > > > > Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
                          > > > > > > objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
                          > > > > > > Nikon CFW 10X occulars with this setup.
                          > > > > > >  
                          > > > > > >  
                          > > > > > >     I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
                          > > > > > > with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars
                          > > > > > with
                          > > > > > > this setup.  
                          > > > > > >  
                          > > > > > >    Please...this is not a 'high-jack', of Rich's thread...please comment
                          > > > > > to
                          > > > > > > Rich (...err, to all)...although we rarely do it...why is not:
                          > > > > > > 'oil-bridgeing the substage-condensor not at least mentioned in this
                          > > > > > > discussion with Rich pondering objectives for his use( okay...Rich already
                          > > > > > > said he is not keen on imerrsion oil useage)?
                          > > > > > >  
                          > > > > > >    Please...as I have a variety of occulars...( okay...I think Rich
                          > > > > > already
                          > > > > > > said he is useing:'10X occulars'.)...please comment on how one need
                          > > > > > concern
                          > > > > > > a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
                          > > > > > > PlanApo?
                          > > > > > >  
                          > > > > > >  
                          > > > > > >     Do occulars even matter in Rich-guys setup?  With the very common: NA
                          > > > > > > 1.25 substage condensors...we often do not 'oil-bridge condensor to
                          > > > > > > slide'..does this factor in to Rich'es 'application Difference'?    thanks
                          > > > > > > for all this practicle/ applied microscopy, charlie guevara     (PS)  in
                          > > > > > > hobby astronomy...Apo refractors are costly, their proud owners seek out
                          > > > > > > planetary observing agendas for crisp/'tack-sharp'  color observations.  
                          > > > > > > Deep-sky (beyond our Suns family of interacting systems)...deep sky
                          > > > > > > observers always 'chant': so little color differences in 'deep sky
                          > > > > > > objects'...no need to worry about: 'purple fringes on bright stars...go
                          > > > > > > 'achro'...leave 'apos' for when you win the lottery.
                          > > > > > >
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > >
                          > > > > > ------------------------------------
                          > > > > >
                          > > > > > Yahoo! Groups Links
                          > > > > >
                          > > > >
                          > > >
                          > >
                          >
                        • Selwyn St Leger
                          Hi Randall, Yes, I was distinguishing between achieving high resolution and being able to use the information it gives. I suggested that in some circumstances
                          Message 12 of 21 , Nov 5, 2011
                            Hi Randall,

                            Yes, I was distinguishing between achieving high resolution and being able to use the information it gives. I suggested that in some circumstances high resolution adds nothing useful because what is resolved has no obvious interpretation.

                            As to the other matters you raised they have been dealt with by others better than I could do.

                            Selwyn


                            --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@...> wrote:
                            >
                            >
                            >
                            > Hi Selwyn,
                            >
                            > Thanks for your comments. Am I correct in paraphrasing, achieving optimum
                            > resolution is not the same as being able to use it in a meaningful way?
                            >
                            > There is another aspect of this subject that I haven't "resolved". Using
                            > very high NA objectives (either dry or oil) with a wet-mount specimen does
                            > not seem to be practical due to what appears to be spherical aberration
                            > induced by the low index (~1.3) water layer under the coverslip. (
                            > Objectives designed for water immersion are the exception, of course. )
                            >
                            > On the other hand, diatom observers often use high index (higher than the
                            > 1.52 of glass or immersion oil) mountant material (index ~2+) with great
                            > success.
                            >
                            > Do you have any thoughts on this matter?
                            >
                            > Best,
                            >
                            > Randall
                            >
                            >
                            >
                            >
                            >
                            >
                            >
                            >
                            > -----Original Message-----
                            > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                            > Behalf Of Selwyn St Leger
                            > Sent: Thursday, November 03, 2011 11:23 AM
                            > To: Microscope@yahoogroups.com
                            > Subject: [Microscope] Re: Application Difference?? 100X PlanApo vs, 63X
                            > Plan-NeoFluar
                            >
                            >
                            > HiRandall,
                            >
                            > Those are interesting observations and well put.
                            >
                            > One point you raise merits further discussion. That is whether or not the
                            > condenser diaphragm should be reduced below NA 1.4 when employing a 1.4 NA
                            > objective.
                            >
                            > With geometric optics the condenser diaphragm is conjugate with (in focus
                            > at) the back focal plane of the objective. Thus, from this way of
                            > considering light transmission its effect is exactly the same as an
                            > equivalent stop in the back focal plane. However, as the image is formed
                            > from diffracted rays off the specimen some of these pass beyond the
                            > geometric light cone, impinge on the periphery of the objective to
                            > contribute to interference at the back focal plane.
                            >
                            > Thus, stopping down the condenser a little does not have a large effect on
                            > resolution. Also, apochromatic objectives (which they must be to attain 1.4
                            > NA) because of their corrections require less stopping down of the condenser
                            > than achromats to achieve good contrast.
                            >
                            > Nevertheless, your broader point has great merit. Attempting to get maximum
                            > resolution is not often, in contexts such as mine, worth the candle. Very
                            > high resolution has to be accompanied by the facility to interpret what has
                            > been resolved. That's fine with test diatoms but looking, say, at living
                            > cytoplasm in a protist there is not much point in being able to perceive a
                            > few more anonymous granules.
                            >
                            > Selwyn
                            >
                            >
                            > --- In Microscope@yahoogroups.com, "Randall Buck" <rbuck@> wrote:
                            > >
                            > >
                            > > Hi,
                            > >
                            > > The more I think about the subject, the more I come to believe that John
                            > W.
                            > > (ajohn) has condensed the whole subject into a very succinct statement:
                            > > (Paraphrasing), Think of Very high NA objectives as though they are
                            > designed
                            > > for one thing only, viewing something that is attached to the underside of
                            > a
                            > > precision coverslip (one that meets the design specifications of the
                            > > objective in hand).
                            > >
                            > > You certainly don't want to be looking through anything other than
                            > immersion
                            > > oil * and the coverslip otherwise you will introduce an amount of
                            > spherical
                            > > aberration that will reduce the performance your expensive objective to
                            > that
                            > > of an objective for which you could have paid much less. Even mountant,
                            > > unless it is more of a filler than an intervening layer may require a
                            > > thinner coverslip to compensate for any added thickness.
                            > >
                            > > Also, remember that in order to take full advantage of the high NA of the
                            > > objective, you will need a high NA condenser. Since the condenser is
                            > > operating at high NA it requires improved optical correction to deliver a
                            > > precisely imaged beam to the subject plane. This is why the aplanatic
                            > > achromat was developed. Of course, it must be oiled to the bottom of the
                            > > slide, in fact, it will generally be impossible to align it of Kohler
                            > > without oiling. In addition, the iris should be used wide open, otherwise
                            > > your expensive 1.4NA aa condenser will be doing the job of a much less
                            > > expensive (lower NA) condenser and taking the performance of your High NA
                            > > objective down with it.
                            > >
                            > > This means that the perceived contrast improvement ordinarily gained by
                            > > reducing the condenser iris to 2/3 cannot be employed when the maximum
                            > > resolution of a high NA objective is the goal. In turn, this means that
                            > > the specimen itself must be of intrinsically high contrast (via staining,
                            > > etc) to be seen at all.
                            > >
                            > > That is not to say that pursuing ultimate resolution isn't worth while or
                            > > fascinating in and of itself but it is a very precise endeavor.
                            > >
                            > > NOTE:
                            > >
                            > > * Older objectives were designed for appropriately aged cedar oil with
                            > each
                            > > lens manufacturer providing their own optimum formulation. The index of
                            > > refraction of cedar oil increases with aging. Newer (does anyone know the
                            > > approximate transition date?) objectives were designed for Shillaber's
                            > (now
                            > > Cargille) non-drying synthetic oils, with a (sodium "D" line index of
                            > > refraction of 1.5150 at 25 deg C).
                            > >
                            > >
                            > > Randall
                            > >
                            > >
                            > >
                            > >
                            > >
                            > > -----Original Message-----
                            > > From: Microscope@yahoogroups.com [mailto:Microscope@yahoogroups.com]On
                            > > Behalf Of Charles Guevara
                            > > Sent: Tuesday, November 01, 2011 7:53 PM
                            > > To: Microscope@yahoogroups.com
                            > > Subject: Re: [Microscope] Application Difference?? 100X PlanApo vs, 63X
                            > > Plan-NeoFluar
                            > >
                            > >
                            > >    Thank you so much, Rich, for this practicle/hands on microscopy thread.
                            > >  
                            > >   Rolf, Randall, David have clarified performance for visual, vrs. image
                            > > capture microscopy with :100X Plan Apo, vrs. 'dry lens' 63X
                            > > Plan-NeoFluar...or 'dry' 63X-60X Plan achro/NA 0.859 ?with correction
                            > collar
                            > > for coverslip/water colum variations?.
                            > >  
                            > >    Rolf noted that the specimen context: (wetmount-pondlife slides with
                            > > coverslips, vrs. flat blood smear, vrs thin tissue section)...for me, Rolf
                            > > noted that this context renders: 'Plan objectives utility as often over
                            > > rated'.  Rolf futher noted ( to my understanding, Rolf!) that for your
                            > > bloodwork observations,Rich,...try for oil-immersion 100X objectives (even
                            > > though you do not like to muddle with oiled-optics...as a 'fall
                            > back...Rolf
                            > > suggested : water immersion/LOMO objectives.).
                            > >  
                            > >    I have : NA 1.25 condensors on both my work-horse stands....I have a
                            > > Nikon Plan 60X,NA 0.85, 160/0.11-0.23 (correction collar objective)...this
                            > > objective functions with a Nikon Phase Turrent/1.25 NA condensor...I have
                            > > Nikon CFW 10X occulars with this setup.
                            > >  
                            > >  
                            > >     I have a Reichert/Austria brightfield /1.25 NA condensor , I enjoy it
                            > > with it's brightfield 100X/1.25 NA oil-objective...I have B10X occulars
                            > with
                            > > this setup.  
                            > >  
                            > >    Please...this is not a 'high-jack', of Rich's thread...please comment
                            > to
                            > > Rich (...err, to all)...although we rarely do it...why is not:
                            > > 'oil-bridgeing the substage-condensor not at least mentioned in this
                            > > discussion with Rich pondering objectives for his use( okay...Rich already
                            > > said he is not keen on imerrsion oil useage)?
                            > >  
                            > >    Please...as I have a variety of occulars...( okay...I think Rich
                            > already
                            > > said he is useing:'10X occulars'.)...please comment on how one need
                            > concern
                            > > a match of: occulars to either a : 63X Plan-NeoFluar, vrs to a : 100X
                            > > PlanApo?
                            > >  
                            > >  
                            > >     Do occulars even matter in Rich-guys setup?  With the very common: NA
                            > > 1.25 substage condensors...we often do not 'oil-bridge condensor to
                            > > slide'..does this factor in to Rich'es 'application Difference'?    thanks
                            > > for all this practicle/ applied microscopy, charlie guevara     (PS)  in
                            > > hobby astronomy...Apo refractors are costly, their proud owners seek out
                            > > planetary observing agendas for crisp/'tack-sharp'  color observations.  
                            > > Deep-sky (beyond our Suns family of interacting systems)...deep sky
                            > > observers always 'chant': so little color differences in 'deep sky
                            > > objects'...no need to worry about: 'purple fringes on bright stars...go
                            > > 'achro'...leave 'apos' for when you win the lottery.
                            > >
                            >
                            >
                            >
                            >
                            > ------------------------------------
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