Zeiss water immersion objective front lens damage

[microcosmos]

I've had a chance to examine the front lens of this Zeiss C-apo 63x/1.2 water immersion objective that was being used on an inverted microscope for life sciences microscopy and possibly cell cultures.

The front lens has a haze that could not be cleaned with 100% ethanol or water:


front lens as seen with coaxial reflected light;


front lens as seen with oblique reflected light.

There is no visible birefringence or bireflectance in either transmitted or reflected polarized light.

The random, evenly distributed pattern on the glass seems to suggest that some kind of aqueous or other solution was left to dry on the lens (not cleaned off after use) and when the solvent evaporated it left a residue.

I can't really tell if the substance has actually eaten into the glass.

Does anyone know what substance could have caused this, and whether the damage is salvageable? What other cleaning agents are safe to try on this particular objective? Note that there appears to be some damage to the lens mount so cleaning agents could seep inside:


(front lens as seen with coaxial reflected light, photographed through 10x eyepiece and 4x objective)

Thank you very much for any ideas.

[Hobbyst46]

Assuming that the dirt (?) is on the outer surface only, I would start with a mild soap solution on a Q-tip, very very gently riping (not rubbing). Then the same with distilled water.
If that fails, I would try the same method but with solvents: isopropanol, then (if it fails) heptane, octane, petrol ether, and if they fail - xylene. In all cases, the same gentle wiping and the minimal amount of solvent.
If all those fail, I would try and check if the dirt is on the inner surface of the lens.

[Scarodactyl]

A higher mag view with coaxial illumination would probably make it a lot easier to tell what is going on. But my bet is that it's been etched.

[tpruuden]

Solidified immersion oil?

[microcosmos]

Thank you all for your suggestions. I forgot to mention that the damage is on the outer surface of the lens - it's not very obvious from the images shown.

I notice that acetone was not suggested as one of the cleaning agents - I assume that's because it could dissolve the cement holding the lens.

We are wondering what substance used in the lab would be able to etch the lens (or its coating?) - so far we can think of no obvious culprits among the reagents usually used in the lab for culturing and mounting the biological material.

Wouldn't solidified immersion oil be cleanable with ethanol still (or not)? As mentioned earlier, ethanol didn't work.

[abednego1995]

Were you guys using Immersol W? If it is, ethanol would work most of the time. My lab uses cleaning fluid EE-3310 from Olympus.

[Hobbyst46]

Thank you all for your suggestions. I forgot to mention that the damage is on the outer surface of the lens - it's not very obvious from the images shown.

I notice that acetone was not suggested as one of the cleaning agents - I assume that's because it could dissolve the cement holding the lens.

We are wondering what substance used in the lab would be able to etch the lens (or its coating?) - so far we can think of no obvious culprits among the reagents usually used in the lab for culturing and mounting the biological material.

Wouldn't solidified immersion oil be cleanable with ethanol still (or not)? As mentioned earlier, ethanol didn't work.

Acetone was not suggested, neither were even stronger solvents (chloroform, for example). These chemicals might indeed dissolve glass cements. And degrade outer surface coatings.

Glass is etched by hydrofluoric acid - not a common stuff in biological/clinical use AFAIK.
Another glass etchant is a strong caustic (pH 13-14) solution. For example, soda caustic, potassion hydroxide in water. Even if the solutions are dilute, overtime they etch glass.

From the photos, though, I imagine that the damage is to the coating, more than to the glass base itself. Just a guess.

As for ethanol (even 96-100%) - in general, it is not an efficient oil/grease remover. Isopropanol (IPA) does a much better job. Heptane, octane, xylene - better yet. Intuitively, the same goes for solidified oils. Acetone is excellent, but I would avoid it from the reasons stated above.

P.S. the pattern of the damage somehow resembles delamination, in a strange way.

[Scarodactyl]

An objective like this is probably not using conventional glass. ED or fluorite glasses might be significantly easier to damage.

[PeteM]

Depending upon what else was dissolved in the water this immersion lens was used with - it could be a mineral deposit. If so, you're not going to touch it with conventional solvents. Can you find out how this objective was used? Can you do closer imaging to see if it's a residue or actual damage to the coating or glass?

The lens looks near unusable as is - it could be that aggressive treatments targeted to whatever that residue (if it's a residue) is would be worth a try. One slight plus - immersion often fills in things like minor damage - might be worth a try as is.

[microcosmos]

Thank you again for all your comments and suggestions. As requested here's a high-magnification micrograph of the lens surface with coaxial reflected light.



The image is white balance-corrected for the halogen light source; sorry for the CA. It seems that the lens surface being examined is slightly off perpendicular to the optical axis so there's a focus shift from left to right - hopefully that has the unintended positive effect of showing more clearly what is happening on the lens surface.

It does look to me as if the coating has been damaged but the glass is largely intact with some scratches?

As far as we know, the lens shouldn't have touched anything other than water, cell culture buffer, and possibly immersion oil (by mistake, as there is an oil objective on the same microscope).

We will try some of the suggested cleaning agents when we get a chance.

[apochronaut]

I have had about 6 objectives with damaged external coatings in my hands and none of them looked like that. Some coatings are vulnerable to certain solvents: ethanol is a prime suspect, isopyopyl not. Usually the coating comes off in complete sections and the resulting image loses contrast slightly.

Objectives typically have a crown glass front lens. This is a norm and in the case of an oil immersion lens that serves to homogenize the immersion system.
A water immersion lens on the other hand cannot be homogeneous. I have seen at least one water immersion objective that used a compound front element. Is it possible that is a cement residue , remaining after a planar front surface departed from it's rearward lens element?

[MichaelG.]

My personal experience of lens-coating damage is limited to crazing [at Kodak] and mould-growth [at BAe] … your excellent image resembles neither of these, so I searched a little wider:

This short paper may be of some interest: https://www.researchgate.net/publicatio ... MIAN_GLASS

… especially the comments about storage

MichaelG.

[apochronaut]

Zeiss had at least one design in their 160mm era that used a moving front correction or distance plate in order to effect an adjustment of the front lens distance to the immersion medium. In other words, the front lens did not contact the immersion medium, just the corrector plate, the adjustment of which allowed for various immersion media.

The front lens itself was compound, comprising a plano convex full width front lens cemented into a concave well which was ground into the back lens of the front doublet.

The immersion contact was against a flat corrector plate, which could be extremely close to the front lens. The reason for this was twofold, I would think.
1) the achromat doublet front lens with a cemented concave interface was no doubt designed to increase planarity, similar to the embedded front lens invented by Arthur Shoemaker. In an embedded design there is no cement, so the front of the objective is free from any deleterious effects of the immersion medium upon cement. In a cemented design, contact with immersion medium is fraught with problems. That's why immersion objectives have front singlets or since Shoemaker's invention, cementless embedded front elements. Having a front corrector plate as the immersion contact surface avoids the cement/immersion medium conflict.
2) A corrector plate would allow for a more refined adjustment of the refractive index at the front of the objective. Water at 1.33 is too low to be homogeneous with glass, so optics up front need to adjust for a high level of s.a in the front elements as well , in this case begin the process of flattening the image. A pretty complex task, right at the front.

[Hobbyst46]

It does look to me as if the coating has been damaged but the glass is largely intact with some scratches?

By chance, I have recently seen damage to mirrors, caused by delamination: photos of minerally coated optical fused silica, where the coating has been delaminated due to various treatments.
The pattern in your photos of the objective lens surface resemble them very much. And IMHO indirectly supports the hypothesis that the coating of the objective lens has been damaged.

[apochronaut]

I could see that, except several of the images depict a surface that has texture, or a degree of depth, almost like a pebbled glass surface. Since coatings are 2 nanometers thick roughly, how would they cause that?

[Hobbyst46]

I could see that, except several of the images depict a surface that has texture, or a degree of depth, almost like a pebbled glass surface. Since coatings are 2 nanometers thick roughly, how would they cause that?

AFAIK the thickness of coating layers is of the order of magnitude of light wavelength - hundreds of nm, so the total coating might be about a micrometer. Info based on reading Wikipedia though...

[apochronaut]

Splitting hairs... Irregardless, how does that potentially create a stippled surface?

[Hobbyst46]

Could be a photo artifact rather than true 3D ? the illumination appears somewhat oblique. In addition to the non-uniform sharpness ?

[Hobbyst46]

As far as we know, the lens shouldn't have touched anything other than water, cell culture buffer, and possibly immersion oil (by mistake, as there is an oil objective on the same microscope).

A common antireflection coating material is Tantalum oxide. Like glass, it is damaged by hydrofluoric acid, but also by caustic alkalies. If anyone was cleaning a biologial uncovered slide with caustic soda while the lens was still in place...stupid, I know...

Another idea, perhaps far-fetched:
The delaminated coatings that I had seen some time ago on a silica mirror. Delamination was caused by heat (a few hundred degrees C, for a short time). If the objective in question has been fitted on a halogen-lit microscope, and if the lamp was turned on for long periods at full power, and if the immersion water had dried/evaporated, (all highly unlikely "and ifs" in series ) then perhaps intense heat had destroyed the objective lens coating. Wild guess.

[apochronaut]

What kind of microscope achieves 200 ° C. at the stage? I wouldn't recommend that you ever apply for a job in the Zeiss marketing dept.

[Hobbyst46]

What kind of microscope achieves 200 ° C. at the stage?

Time and temperature are sometimes (not always) equivalent. I never claimed 200°C, I was guessing 60°C (maybe). Would be interesting to know the long-term stability of a microscope lens coating under (hypothetical) focussed halogen illumination.

I wouldn't recommend...

Why so ???

[apochronaut]

The wording was " a few hundred degrees C, for a short time " : that's even more than two hundred. Zeiss might not want it known that it was possible to melt solder on a biological microscope stage, with just their on board halogen illuminator.

[tpruuden]

Try acetone/IPA mix and relatively aggressive scrape with sharpened wooden stick on one edge - if it is residue, it starts to move and there is not much to lose.

[viktor j nilsson]

On one objective, I had a really stubborn layer of dried up immersion oil. It literally took a hundred q-tips with various solvents before I got everything off. And it took quite many attempts before I started to be able to get it loosened up. So it's worth trying a bit more with different solvents before you give up.

[Sure Squintsalot]

Assuming it's a glass lens, I'd try white vinegar on a Qtip, since organic solvents did nothing.

If you really have nothing to lose with this objective, your next step might a very light repolishing with 0.3 micron aluminum oxide compound. Make a wax cast of the lens to use as your form, put it on the end of a round toothpick, and load it up with polishing compound, then some gentle spinning over the lens surface until it's smooth. An hours work may yet save this.

[toomanyhandles]

Tissue culture growth media have salts of course but especially, proteins (think dilute blood plasma or fetal bovine serum). If dilute enough you may not get the opaque results as it dried/ aged as one may expect.

You may want to look at solvents useful for proteinaceous materials.

[microcosmos]

Here are photos of the reflected light setup that I used to micrograph the front lens of the objective.


To create enough vertical space for the objective, I removed the stage insert and transmitted light condenser from my Olympus BHSP and placed the stage insert and the objective on the condenser rack.


The imaging objective is an MSPlan 50x. Despite using the original Olympus BH2-MA-2 tube lens and the original NFK photo eyepiece that were designed to compensate for the objective aberrations, I still get CA (see earlier micrograph). This could be because of the additional tube length and lens in the BHSP analyser attachment (not shown) that sits on top of my epi-illuminator. The objectives are covered in gaffer tape to hide the modern printed Microsoft Word Arial-type font that drives me crazy.

[microcosmos]

Thank you everyone for your many suggestions and the fascinating insights into non-homogeneous immersion objective design and glass disease.


I have tried multiple applications of isopropanol and heptane with lens paper but both of them seem to have no effect at all, judging from assessments with a 10x loupe. I did not have cotton buds handy at the time.

I will move on to other solvents next week.

[microcosmos]

Acetone and chloroform both had no visible effect under a 10x loupe.