MicrobeHunter.com Microscopy Forum

Planapo 63x/1.4, DIC, 155µm length - 8µm width, stacked and stitched in Photoshop

This is from a Klaus Kemp 8 Form slide

Hi Glen,
nice image! This is probably one of the diatoms that end up in broken pieces in many cases after cleaning.

Bob

Thats really good! Excellent contrast and resolution, thanks for sharing.

Thanks MicroBob,

I am afaid I can't take any credit for its condition as I had nothing to do with its cleaning or mounting.

It is from a Klaus Kemp 8 Form Test slide

Here is some info on the diatom: https://diatoms.org/species/nitzschia_sigma1
.

Thanks Wes

Out of curiosity do you see the striae in Amphipleura pellucida with the 63x planapo? It might be the right subject to also test the effect of condenser oiling on resolution.

Wes wrote: ↑

Sat Nov 23, 2019 5:52 pm

Out of curiosity do you see the striae in Amphipleura pellucida with the 63x planapo? It might be the right subject to also test the effect of condenser oiling on resolution.

Funny you should ask, had a look today while I was imaging the Nitzschia sigma but had no luck.

I seem to recall being able to resolve the puncta previously but did not manage to do so today.

Which leads me to suspect that it requires a more subtle setup.

Nicely done. Thanks for sharing it.

Great Photos 75RR!

Maybe you need to try one of the older Zeiss Objectives with n.A. of incredible 1.6:
PlanApo 100/1,6 Imm or the 74/1,6

-
Thanks PeteM

Maybe you need to try one of the older Zeiss Objectives with n.A. of incredible 1.6:
PlanApo 100/1,6 Imm or the 74/1,6

Love to try, though I fear I would not be able to do them justice

-
Thanks PeteM

Maybe you need to try one of the older Zeiss Objectives with n.A. of incredible 1.6:
PlanApo 100/1,6 Imm or the 74/1,6

Love to try, though I fear I would not be able to do them justice

ImperatorRex wrote: ↑

Sat Nov 23, 2019 9:05 pm

Great Photos 75RR!

Maybe you need to try one of the older Zeiss Objectives with n.A. of incredible 1.6:
PlanApo 100/1,6 Imm or the 74/1,6

NA of 1,6?? I had never heard of that till now. I know some of the modern 100x planapos used for TIRF microscopy deliver NA of 1,46. Can you provide a bit more info, this sounds remarkable.

Wes wrote: ↑

Sat Nov 23, 2019 10:03 pm

NA of 1,6?? I had never heard of that till now. I know some of the modern 100x planapos used for TIRF microscopy deliver NA of 1,46. Can you provide a bit more info, this sounds remarkable.

I assumed that those were the objectives used in the famous 1888 catalogue ... turns out they used a 1.3NA

This is the blurb under the image as translated by google:

Pleurosigma angulatum. Enlargement: 4900.
Objective: Apochromat 2.0 mm, 1.30 aperture; Projectios Ocular 4.
Illumination: central, solar image projected into the object plane by Apochromat 16.0 mm, 0.30 Apert.

Here is the link: https://ia800202.us.archive.org/9/items ... 109582.pdf

Which leaves me with the same question as yours

Found the link!

"All these photos were taken with the Zeiss lens 1/10 inch, nA 1.63. Monochromatic sunlight. Special compensation eyepiece 12. Condenser 1.6. Inclusion in a medium with refractive index 2.4. Coverslips and flint glass slides with a refractive index of 1.72."

Image is in the post above the one I linked to

viewtopic.php?f=15&t=6806&p#p60731

Thanks 75RR for the post of the 4900x enlargement. It is impressive, and as previously discussed, was achieved by means of illumination, condenser, media and slide optimization; likely a 1.6NA objective alone would not provide such nice details.

Hobbyst46 wrote: ↑

Sun Nov 24, 2019 8:39 am

Thanks 75RR for the post of the 4900x enlargement. It is impressive, and as previously discussed, was achieved by means of illumination, condenser, media and slide optimization; likely a 1.6NA objective alone would not provide such nice details.

I agree ... and I think our inability to match such results is, in part, attributable to the scale of our image sensors.
That image was [if I understand correctly] produced on the equipment shown in plates XII and XIII.

There was recently a rather fruitless discussion here, about pixel size and ‘resolution’
See message #15 et seq here: viewtopic.php?f=9&t=5907#p72498

That Zeiss equipment is using a very large “sensor” ... so the grain in the emulsion is providing the equivalent of very small pixels. Q.E.D.

MichaelG.

Regarding the 4900x image : It is interesting to speculate how good the original silver image must have been.
It is very clear that the image we are seeing has been considerably degraded by the scanning process ... and the published image was, of course, already poorer than the original.

MichaelG.

75RR wrote: ↑

Sun Nov 24, 2019 7:09 am

Found the link!

"All these photos were taken with the Zeiss lens 1/10 inch, nA 1.63. Monochromatic sunlight. Special compensation eyepiece 12. Condenser 1.6. Inclusion in a medium with refractive index 2.4. Coverslips and flint glass slides with a refractive index of 1.72."

Image is in the post above the one I linked to

viewtopic.php?f=15&t=6806&p#p60731

Thanks!

Its impressive to say the least, especially considering the time when it was realized.

Hi together,
as far as I know these Zeiss n.a. 1,6 objectives were used with a different immersion medium and had very fragile front lens mounting. The n.a. 2,4 mountant must have been arsenic yellow medium. These objectives were probably available until the TEM and SEM became available. Today it would probably be possible to go beyond n.a. 1,6 in light microscope objective and condensers but nobody cares. What is used in confocal microscopy?

Bob

MicroBob wrote: ↑

Sun Nov 24, 2019 10:15 pm

Hi together,
as far as I know these Zeiss n.a. 1,6 objectives were used with a different immersion medium and had very fragile front lens mounting. The n.a. 2,4 mountant must have been arsenic yellow medium. These objectives were probably available until the TEM and SEM became available. Today it would probably be possible to go beyond n.a. 1,6 in light microscope objective and condensers but nobody cares. What is used in confocal microscopy

Bob, in "simple" confocal microscopy the resolution is still described in terms of the Abbe or Raleigh rules. And the same objectives are used (say, NA up to about 1.4, mostly). The point is, that a high NA is favored in confocal because it gathers more light; and confocal is essentially fluorescence microscopy. The basic advantage of confocal is removal of out of focus images.
There are more sophisticated, super-resolution confocal microscopes, where resolution is not limited by diffraction; however, as much as I remember, super-resolution is achieved not by particularly high NAs but by other means..