Dodging the infinity hump

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Beatsy
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Re: Dodging the infinity hump

Post by Beatsy »

Chris S. wrote:
Mon Mar 07, 2022 11:11 pm
... Condenser mounting is an interesting question--maybe put it on a second, shorter BiSlide? Or as a distant second thought, could the condenser perhaps be mounted on a second carriage on a single BiSlide?

Fun stuff to think about.

--Chris S.
Certainly an interesting project, but thoughts of the condenser was the last straw that put me off the idea. I forgot to mention that. You need really accurate Koehler illumination setup at the highest resolutions, and suitable controls to achieve it. I felt I'd be recreating so many parts of a standard microscope frame, it might just as well be a standard microscope frame. That, and already having too many open projects on the go made it a much less attractive proposition in the end.

It's not totally off the table, but I see it more as a Plan B or C now, if the other routes go pear shaped. I'll have more of the required components in hand if/when that happened too...

Thanks for your thoughts though - alternative perspectives always welcome.

Scarodactyl
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Re: Dodging the infinity hump

Post by Scarodactyl »

Yeah, it seems like it would be cheaper and less frustrating to wait until a research stand comes up for sale cheap. A few months back I got a Nikon E800 very affordably, including a DIC nosepiece, condenser and some prisms. And of course I already had a 20x planapo for it thanks to eBay.
Of course that is neither here nor there for trying shorter wavelengths, which does seem like a smart approach.

Alan Wood
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Re: Dodging the infinity hump

Post by Alan Wood »

You might want to check out this blog by Quekett member Jonathan Crowther, currently looking at diatoms in UV:

https://jmcscientificconsulting.com/blog/

Alan Wood

Beatsy
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Re: Dodging the infinity hump

Post by Beatsy »

Alan Wood wrote:
Tue Mar 08, 2022 2:37 am
You might want to check out this blog by Quekett member Jonathan Crowther, currently looking at diatoms in UV:

https://jmcscientificconsulting.com/blog/

Alan Wood
Co-incidentally, it was recent correspondence and conversations with Jonathan (ref diatoms) that prompted me to explore this route. He also has to use very short wavelengths in his work though, so it's all quartz slides and coverslips, with a limited selection of not-so-high NA objectives. All mountants are practically opaque at <300nm too. A bit too extreme for my needs, but still very interesting.

Beatsy
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Re: Dodging the infinity hump

Post by Beatsy »

Beatsy wrote:
Mon Mar 07, 2022 3:04 pm
Scarodactyl wrote:
Mon Mar 07, 2022 1:22 pm
If you ever get an SIL objective working you will be a true hero. I think any enthusiast who has run across them has been curious what those marvelous NAs might reveal, but so far nobody's had the gumption as far as I know. I sure haven't anyway.
Won't be me. The RI of the substrate (which would be the mountant in/around a diatom in my case) has to be at least as high as that of the SIL material or NA is limited to the RI of the substrate. Tough call to match the 2.35 of the lens linked! Probably meant for imaging deeper metal layers through silicon on ICs, or something related to that (RI of silicon is 3.4).

Zrax, a high-RI diatom mountant has an RI of 1.7+ when properly hardened. This is a good match for glass and sapphire SILs (hemispheres). There is a better shape for the SIL too, the super-sphere, which is simply a bit more than half a sphere, say 3/5ths. This increases NA by up to RI squared :shock: I can envisage a way of starting with a sphere and grinding/polishing a flat like that.

This is a very intriguing little rabbit hole that I'm going to continue exploring anyway - independently of the UV stuff.
Just a bit of teaser at this point as this is a very fragmented project that I'm only visiting sporadically - but I hit a milestone. Since the idea started here, it seemed the best place to put this (one-off) report.

I do not claim to have made proper SuperSILs here (Super Solid Immersion Lenses) but they're definitely good "working models" of SuperSILs. I need to learn more about handling during fabrication and in-use before spending serious money on real spherical lenses to make SuperSILs from - which is why I'm going the cheap and crude route for now. After a few false starts trying to make spheres (beads) from scratch I found some low/mid grade glass ball bearings for pennies each and bought a selection of sizes from 1mm-3mm. Easier to make glass spheres if you start with glass spheres :D

I fire polish these (chefs blowtorch on ceramic surface), pick out the clear spherical successes (about 1 in 2 or 3 is "good enough), glue 4 to a glass slide (to keep them level when lapping), grind and polish the flat on ascending-grit lapping papers stuck to glass (sphere diameter to height ratio dependent on RI), then cook them out of the glue. Quick clean and they're done.

These are 2mm diameter SSILs. A bit grubby from the glue and far from optically perfect, but they actually work surprisingly well under visual inspection (no photos yet). I placed one "dry", flat side down, on various bits of an open diatom strew and was immediately able to see far more detail in the magnified images through the SIL than I could just looking directly at the diatoms - even if I added higher power oculars for extra magnification with the scope alone. Resolution was subjectively better, I'm convinced of it, but I fully intend to photograph and measure the improvement objectively once the process is less manual and fiddly. I have spare manipulator parts laying around that should help improve that.
SuperSILs 2mm.jpg
So although this still looks very promising I'm not yet ready to spend >£70 each on optically perfect, high-RI, 1mm spheres to turn into SSILs. I intend to try more of these home made ones to find the best size for my needs first (diatoms) and the practicality in use. The diatoms in my early tests had to be squashed flat (WD<50nm in air) but I think mountant will improve working distance by 200nm or so. I need to be more accurate with the grinding too as I usually only get one of the four that is exactly the right height (+/- 10 microns). That's partly because of size variation in the source balls, but partly technique too. I welcome the variation for experimenting, but once I find the sweet spot, I want to hit it for all the spheres being "modified" thereafter. Which means I have quite a bit more sporadic tinkering to do before I'll have anything else worthwhile to report. That should be with visual results (pics) too - but I'll start a new thread for that...

Lou Jost
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Re: Dodging the infinity hump

Post by Lou Jost »

This is a new level of DYI project! This is going to be fascinating to watch from a distance. I hope it works out for you.

Macro_Cosmos
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Re: Dodging the infinity hump

Post by Macro_Cosmos »

Mounting diatoms in arsenic trisulfide with a high refractive index (sapphire!) coverslip utilised could be a fun route to explore as well: https://en.wikipedia.org/wiki/Arsenic_trisulfide
Colloquially known as "realgar", refractive indices up to 2.45.

rjlittlefield
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Re: Dodging the infinity hump

Post by rjlittlefield »

I'm still scratching my head a little, trying to get solid on how all this fits together.

My understanding of the SIL process is that
(A) the thing to be imaged is embedded in high RI material (or at least is close enough to the high RI material that near-field coupling works), and
(B) the shape of the lens is such that light is collected from the subject at a wide range of angles.

As a result, the final image benefits from two aspects: the short wavelength of light in the high RI material, combined with the wide range of angles that contribute to the image.

Viewed in those terms, the SIL is nothing more (and nothing less!) than the front element of an immersion objective that is optimized for use with high-RI mountant. The ultimate resolution can theoretically be as high as NA = RI of the mountant, in the limiting case that the lens collects light over a full 180 degrees, but it will never get any higher than that.

It's always possible that I've missed something, but I think the advertised benefit of a factor of RI squared applies only when the comparison is made against the dry NA of the rear optics that pair with the SIL to comprise the whole imaging system. That is a benefit, but perhaps it should be thought of as implying that you can get by with relatively low NA in the rear optics, and still potentially get the full NA = RI in terms of resolution on subject.

Have I messed this up?

--Rik

Beatsy
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Re: Dodging the infinity hump

Post by Beatsy »

Macro_Cosmos wrote:
Mon Apr 04, 2022 7:35 pm
Mounting diatoms in arsenic trisulfide with a high refractive index (sapphire!) coverslip utilised could be a fun route to explore as well: https://en.wikipedia.org/wiki/Arsenic_trisulfide
Colloquially known as "realgar", refractive indices up to 2.45.
Yeah, I've looked into that, but it involves subliming the realgar onto the mounted diatoms. Not for the amateur IMO (although it always was back in the day). The high RI in this case gives the benefit of more contrast due to the RI difference between specimen (diatoms) and mountant. Resolution comes from the lenses.

Beatsy
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Re: Dodging the infinity hump

Post by Beatsy »

rjlittlefield wrote:
Mon Apr 04, 2022 9:54 pm
I'm still scratching my head a little, trying to get solid on how all this fits together.

<snip>
It's always possible that I've missed something, but I think the advertised benefit of a factor of RI squared applies only when the comparison is made against the dry NA of the rear optics that pair with the SIL to comprise the whole imaging system. That is a benefit, but perhaps it should be thought of as implying that you can get by with relatively low NA in the rear optics, and still potentially get the full NA = RI in terms of resolution on subject.

Have I messed this up?
No, you have it exactly right. For all intents and purposes, the SIL is vaguely like a contact version of a Leeuwenhoek microscope lens. The objective just captures the magnified image from that. But, because the image is pre-magnified by the SIL, at a higher NA than the objective, features that couldn't be resolved with the objective alone are scaled up so it can resolve them easily. The limiting NA of objective isn't a factor in the pre-magnification.

It's still a useful approach in the context of limited access to costly, super-high NA objectives. How useful remains to be determined...

One other thing is keeping me curious. At the near field interface, I've read about an "induced polarisation" effect which is exquisitely sensitive to the distance between the subject and the flat SIL surface. It works in air for sure, and I think it will still work in mountant. Not clear what the range of the effect is yet, but appears to be in the half-wavelength region (say 0-200 nm). It could be useful for extra contrast in brightfield illumination (by viewing through a polariser), so I at least want to continue to a position where I can test that too.

Edit: I am aware that all this near-field/contrast stuff will require dissection of diatom frustules to get small, flat "areas of interest" (e.g. rows of closely spaced pores) close enough to the SIL to show any effects. But I'm already geared up for that, and well practiced at it (for other reasons). So fortunately there appears to be no major barriers to progressing this (for me).

rjlittlefield
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Re: Dodging the infinity hump

Post by rjlittlefield »

Excellent -- thanks for the confirmation and the analogy with a Leeuwenhoek microscope lens.

I'm looking forward to more results!

--Rik

Macro_Cosmos
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Re: Dodging the infinity hump

Post by Macro_Cosmos »

Beatsy wrote:
Mon Apr 04, 2022 11:23 pm
Yeah, I've looked into that, but it involves subliming the realgar onto the mounted diatoms. Not for the amateur IMO (although it always was back in the day). The high RI in this case gives the benefit of more contrast due to the RI difference between specimen (diatoms) and mountant. Resolution comes from the lenses.
We have definitely grown to me more cautious, or "wimpy" as the older generation will put it. "Kids nowadays are scared of hydrochloric acid, back in my days..." The mere thought of boiling acids to clean the frustules spook me, let alone touching arsenic. I have a friend that made some kind of resin-like compound mountant with 2.1 RI -- extreme toxicity. I will see if he has the time to provide me with a slide.

That said, I keep seeing SIL and thinking about silicone oil immersion lenses which are also marked with (lower case) "Sil". What a funny coincidence.

Beatsy
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Re: Dodging the infinity hump

Post by Beatsy »

Beatsy wrote:
Mon Mar 07, 2022 4:21 am
<snip>
Which leaves just one option, I think. Build a 365nm UV capability around the existing Zeiss scope - which is what I now plan to do.
<snip>
Hah!

Time passed. Nothing progressed. Then the plan flexed somewhat. Well - flexed completely really. Dodging the infinity hump now involves wholly embracing it! :D

I bought an Olympus BX61 kitted out for fluorescence (all 6 filter cubes in the turret are populated). Not because I particularly want to do fluorescence, but because the entire light path, including the objectives, transmits well at UV wavelengths down to <365nm. Furthermore, two of the objectives fitted are corrected for soft-UV to near-IR wavelengths and are of renowned high quality (UPlanSApo 100x/1.40 Oil and PlanApo N 60x/1.42 Oil). Given what I paid (private sale) and the going 2nd-hand rate for these two PlanAPOs, I feel like I got the two objectives at a high-but-fair price - with a fully functional BX61 frame and fluorescence accessories thrown in for free! I'm happy.

I don't like it being motorised though; a manual BX51 frame would have been preferred. But all the required control boxes and cables are present and operational, and this system was available now. It also came with near-perfect-for-my-needs PlanAPOs plus 10x and 20x UPlanFL N objectives fitted. So I compromised and grabbed it - and I'm glad I did. The motorised parts have since grown on me for some operations too. Not much, but a little bit.

The optics are a revelation though! The improvement over my beloved Zeiss ICM405 is astonishing. Jaw-dropping. Somewhat upsetting too - when I think how long I've "managed" with the (now) clearly-lesser capabilities of the Zeiss. Oh well, got there in the end :)

I'm awaiting a part to adapt my NA1.4 Heine condenser, but here's one of the first test "snaps" taken with the 100x 1.4 PlanAPO and a BH2 condenser (NA1.4) both oiled to the slide. Straight brightfield, white light, single shot. Contrast stretched in post. The striae are well resolved (about 0.25 micron spacing) but punctae far less so (about 0.22 micron spacing). But this is just a no-effort snapshot in white light that already surpasses the best I ever managed using a PlanAPO 63/1.4 and blue light on the Zeiss. Gear matters!
ap-pmn.jpg
100% crop into the image above...
ap-121.jpg

chris_ma
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Re: Dodging the infinity hump

Post by chris_ma »

I hardly understand anything about the tech, but I always enjoy a good gear story, and I really like the test shot - thanks for sharing!
chris

Macro_Cosmos
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Re: Dodging the infinity hump

Post by Macro_Cosmos »

Which adaptor part did you get? If it is the one on eBay for BH, it will not work on BX. The dovetail clearance is too little for it to be mounted. I designed one specifically for Olympus infinity frames, and it should work on Nikon frames as well.

Here is an example of Heine phase contrast with a Pleurosigma sp. diatom.
Image

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