Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

Here you can post pictures and videos to show others.
Message
Author
MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#1 Post by MicroBob » Fri Aug 09, 2019 7:54 pm

Hi together,

my lab/office renovation is coming to an end. Today I planed the oak window sills and installed them. Only three boards, but difficult ones!

Hobbyst46-Doron has sent me a couple of diatom- and radiolaria slides he has made himself. I find it always a big joy to look through strew slides because there is so much to find it you look long enough.
These diatoms are from the Mediterranean Sea, Israel, and the speciality of the slide is that Doron unsed alternative cleaning and mounting techniques to make it. They were cleaned in SDS, EDTA and H2O2 and mounted in NOA61.

I took pictures of two of the stronger diatoms with the 100:1 Zeiss plan oil objective. The bigger one (39µ, a Scoliopleura?) was cleaned but not separated in the cleaning process and I stacked right through it. But here I show just one image from the upper halve as the stack wasn't able to show the whole structure clearly enough. The second diatom (Cocconeis?) was quite small (about 19µ) but from the contrasty image it has to be quite strongly built.

Pictures with DIC, first one single shot, second one stacked with PICOLAY.

Thank you for the nice slides, Doron!

Bob
Attachments
IMGP0874-RGB bearbeitet 1024.jpg
IMGP0874-RGB bearbeitet 1024.jpg (411.06 KiB) Viewed 1063 times
diatom medt sea Israel stack-RGB.jpg
diatom medt sea Israel stack-RGB.jpg (174.06 KiB) Viewed 1063 times

User avatar
75RR
Posts: 6063
Joined: Sun Oct 12, 2014 2:34 am
Location: Estepona

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#2 Post by 75RR » Fri Aug 09, 2019 8:08 pm

Excellent captures, particularly given the very small size of the diatoms.
They were cleaned in SDS, EDTA and H2O2 and mounted in NOA61.
How do you find the mountant compared to others you have seen?
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#3 Post by MicroBob » Fri Aug 09, 2019 8:18 pm

Doron has sent me a slide made with Pleurax too, so I will be able to compare and report.
NOA61 has a comparatively low r.i. for a diatom mountant and might be better for stronger built diatoms than for finely built ones like Amphipleura pellucida.

MichaelG.
Posts: 1397
Joined: Mon Apr 10, 2017 8:24 am
Location: NorthWest England

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#4 Post by MichaelG. » Fri Aug 09, 2019 8:34 pm

Great images, Bob

I particularly like the little (Cocconeis?), which I don't thinkI have ever seen before.

MichaelG.
Too many 'projects'

Hobbyst46
Posts: 2031
Joined: Mon Aug 21, 2017 9:02 pm

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#5 Post by Hobbyst46 » Fri Aug 09, 2019 9:20 pm

Hi Bob,
Thanks for taking the photos and for posting.
My tentative identification for those diatoms are Petroneis humerosa (the top one, with somewhat pointed ends) and Cocconeis (the oval form). Those were plentiful in the specimen I scooped from the front shallow water line.
Mild cleaning offers less cleavage of the diatom into valves, which is advantage or disadvantage, depending on the purpose of the beholder...
Zeiss Standard GFL+Canon EOS-M10, Olympus VMZ stereo

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#6 Post by MicroBob » Sun Aug 11, 2019 8:28 pm

Hi together,

some more images: Diatoms and a sponge needle, this case mounted in Pleurax.
Leitz NPL Fluotar 50:1 1,0 Zeiss DIC old on 1960s Phomi.
Both stacked with PICOLAY, image editing with GIMP

Bob
Attachments
pysharp_sup0_al1-RGB 1024.jpg
pysharp_sup0_al1-RGB 1024.jpg (130.99 KiB) Viewed 976 times
Schwammnadel.jpg
Schwammnadel.jpg (153.94 KiB) Viewed 976 times

User avatar
75RR
Posts: 6063
Joined: Sun Oct 12, 2014 2:34 am
Location: Estepona

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#7 Post by 75RR » Sun Aug 11, 2019 8:38 pm

Very nice detail on the diatom image!
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

mintakax
Posts: 73
Joined: Mon Jul 29, 2019 12:06 am
Location: Boulder CO, USA

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#8 Post by mintakax » Mon Aug 12, 2019 2:32 pm

Amazing detail ! Thanks

Hobbyst46
Posts: 2031
Joined: Mon Aug 21, 2017 9:02 pm

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#9 Post by Hobbyst46 » Mon Aug 12, 2019 3:33 pm

Hi Bob, thanks again for posting these images!

I guess that the DIC improves them more than does the mountant. BTW, one disadvantage of NOA61 is that sometimes, it does not fully penetrate the frustule, so an ugly air bubble is seen inside the diatom. Warming the NOA61 does not help. NOA61, on the other hand is safer to use, since Pleurax gives off nasty odors of phenol and sulfides (naturally so, they are the building blocks of the stuff).

One can see that the gentle cleaning of the diatoms has preserved intact the frustule, at least for many of the forms.
Zeiss Standard GFL+Canon EOS-M10, Olympus VMZ stereo

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#10 Post by MicroBob » Mon Aug 12, 2019 5:49 pm

Hi Doron,
I was using oil immersion, top and bottom and this made a direct comparison of the slides with NOA61 and Pleurax difficult.
My impression is that Pleurax penetrates very well, but the NOA 61 slides I got from you don't at all show an uncommon amount of unpenetated diatoms.
When comparing the results to that of other microscopists it has to be kept in mind that they usually work with diatom halves only and that they don't show the problem areas of the slide. 8-)
LOCA TP 2500 works great in this respect but has too little r.i. for diatoms.

Your cleaning method does it's job and the diatoms look really clean. So for the amateur there is little reason to use dangerous chemicals at his kitchen table with the children and pets running around.

My Pleurax is from Rosenfeldt's original retry of the old recipe and it is not as clear in colour as the dutch Pleurax. I think I will order some there - it is good to support people who make the effort to cook mountants too.

I have a bag full of "EDTA" but I don't know which variant it is. Does your EDTA solve readily in cold water?

Bob

Wes
Posts: 198
Joined: Sat Mar 09, 2019 12:58 pm

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#11 Post by Wes » Mon Aug 12, 2019 6:15 pm

Great images Bob, thanks for sharing them and the cleaning recipe.
MicroBob wrote: I have a bag full of "EDTA" but I don't know which variant it is. Does your EDTA solve readily in cold water?
Regular EDTA will not dissolve readily in water. You need to add sodium hydroxide which would subtract the protons from EDTA carboxylic groups and in turn yield soluble EDTA anions. A standard recipe is to prepare 0.5 M EDTA at pH 8.0 with the help of NaOH solution.

Hobbyst46
Posts: 2031
Joined: Mon Aug 21, 2017 9:02 pm

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#12 Post by Hobbyst46 » Mon Aug 12, 2019 6:37 pm

MicroBob wrote:I have a bag full of "EDTA" but I don't know which variant it is. Does your EDTA solve readily in cold water?
Normally, "EDTA" in a commercial package might mean either real EDTA (acid), or EDTA disodium salt, aka Na2EDTA. Both are white coarse powders. What I used was a 0.1M solution. That means 37% (w/v), based on Na2EDTA. But I am sure that the exact concentration is not critical.
Note: in the following, "water " means DW!!

EDTA acid does not dissolve in water - cold or hot. If this is what you have, you can dissolve it in a concentrated solution of NaOH - caustic soda (10-20% in water). Prepare and handle the NaOH with great care - don protective glasses/goggles, rubber gloves, shoes and a plastic apron. Still, NaOH is often sold over the counter, as a drain-cleaner, to break stubborn clogs.
Weigh out 28-29g of EDTA into 50ml of water in a glass jar or good quality plastic jar. Add the caustic dropwise, with stirring, slowly (expect some heat release), until the powder disappears. The pH at this stage will be 6-7 (check with indicator paper). Top up the solution with water until the volume is 100ml. Close the jar hermetically when not in use. It keeps forever (no need to fridge) if properly sealed against entry of CO2.

Na2EDTA is much more soluble than EDTA, but not nearly enough for our purpose. If this is what you have, start as before, with 37g of powder this time (instead of 28g). Proceed with the caustic as above. Reach solubility and correct pH. Top up with water as above. The final product is the same as above.

AFAIK, heating is not the way to go in this case.
Zeiss Standard GFL+Canon EOS-M10, Olympus VMZ stereo

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#13 Post by MicroBob » Mon Aug 12, 2019 7:58 pm

Hi Wes and Doron,
thank you for your help indentifying my EDTA. It has to be Na2EDTA as it was soluble in certain amounts in water with a little NaOH added.
My expectation was that the EDTA will bring certain undissolvable elements in my sample into solution. Here the question is, how much EDTA concentration is needed for this purpose? It is probably also important to make sure the EDTA stays in solution until removed or it might contaminate the sample with EDTA crystals. I have enclosed another image from your Pleurax slide, Doron. It shows crystallic particles that seem to be birefringent. They were mainly at the edge of the slide. Is it possible that this is a residue from the cleaning agents, or would you say it is sand?

Bob
Attachments
Crystals at border of slide.jpg
Crystals at border of slide.jpg (157.86 KiB) Viewed 893 times

Hobbyst46
Posts: 2031
Joined: Mon Aug 21, 2017 9:02 pm

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#14 Post by Hobbyst46 » Mon Aug 12, 2019 8:56 pm

MicroBob wrote:My expectation was that the EDTA will bring certain undissolvable elements in my sample into solution. Here the question is, how much EDTA concentration is needed for this purpose?
The main purpose of EDTA, as I understand, it is to solubilize minerals, since it strongly binds metal ions such as the abundant iron[III] or calcium[II] or magnesium[II] ions, in a process named complexation. So ions will be leached out of non-diatom slime. It is difficult to tailor a specific strength of the EDTA (or other chemicals used for cleaning diatoms!!) to the slime in question, since we know nothing about the concentration of minerals in the slime. So, I suggested 0.1M on the basis of the experience of the authors of that research article; and I applied EDTA twice in some cases. BTW, 0.1M is a huge concentration of EDTA in most cases!! A third treatment with EDTA contributed nothing.
It is probably also important to make sure the EDTA stays in solution until removed or it might contaminate the sample with EDTA crystals.
That is right, I rinse the diatoms in a centrifuge tube 8 to 10 times in a row with DW after the chemical cleaning, each time the dilution ratio is 1:10-1:20. This is no guarantee that EDTA was eliminated. A very rough calculation shows that the dried diatom-coated coverlip might carry up to around 20ng (nano-grams) of EDTA. And given the possible weight of the birefringent "crystals" in the image as 1ng, the possibility that these are EDTA cannot be ruled out.
BTW, the same suspicions apply to any chemical in the diatom-water mixture.
HOWEVER, my answer to the next question:
I have enclosed another image from your Pleurax slide, Doron. It shows crystallic particles that seem to be birefringent. They were mainly at the edge of the slide. Is it possible that this is a residue from the cleaning agents, or would you say it is sand?
I believe they are simply sand, quartz particles. I do not know if Na2EDTA or EDTA crystals are so strongly birefringent as those in the photo. I do not know their form, if they are cubes, polyhedra or needle-like. Furthermore, I doubt that EDTA crystals would withstand the boiling-hot Pleurax mounting...
But that is no proof either...
Zeiss Standard GFL+Canon EOS-M10, Olympus VMZ stereo

User avatar
75RR
Posts: 6063
Joined: Sun Oct 12, 2014 2:34 am
Location: Estepona

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#15 Post by 75RR » Wed Aug 14, 2019 12:49 pm

Hobbyst46 wrote: Note: in the following, "water " means DW!!
Just a heads up on something we are all guilty of. Acronyms!

We forget sometimes that many of the people visiting the forum and indeed some new members are newbies ...


For example, there is nothing more ubiquitous than objective and condenser NA, yet finding out what it means for a beginner is not as straight forward as one might assume, and therefore in many cases is off-putting.

So, not to contribute to the headaches ... NA stands for Numerical Aperture:

"The numerical aperture of a microscope objective is a measure of its ability to gather light and resolve fine specimen detail at a fixed object distance."


https://www.microscopyu.com/microscopy- ... l-aperture

In a nutshell, the higher the better. Yet as in most things in microscopy (and life) everything comes at a cost. (i.e. more light is needed and a there is shallower depth of field for example)


As for the original quote - I am going to assume the DW means Distilled Water - hope I am right! :)
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#16 Post by MicroBob » Wed Aug 14, 2019 12:53 pm

75RR wrote: I am going to assume the DW means Distilled Water - hope I am right! :)
Oh no - I just tried it with diluted wiskey! :roll:

User avatar
75RR
Posts: 6063
Joined: Sun Oct 12, 2014 2:34 am
Location: Estepona

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#17 Post by 75RR » Wed Aug 14, 2019 1:05 pm

MicroBob wrote:
75RR wrote: I am going to assume the DW means Distilled Water - hope I am right! :)
Oh no - I just tried it with diluted wiskey! :roll:
"Eye-rolling has been present in literature since at least the 16th century, according to the Oxford English Dictionary.[2] William Shakespeare periodically would use the gesture in his works to portray lust or passion for another character, as used in his poem The Rape of Lucrece.[1][3] In this time period, eye-rolling was used commonly as an expression of desire or flirtation, and it continued to be used this way in literature for centuries. Up until about the 1950s this same meaning was used in music and films, but began translating to the meaning known today. The widespread use of eye-rolling in a negative connotation wasn't present until the 1980s.[4]"

https://en.wikipedia.org/wiki/Eye-rolling

I suppose this begs the question: are you over forty or under forty, and if over should I worry? ;)
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#18 Post by MicroBob » Wed Aug 14, 2019 1:32 pm

Clearly over and in love with the improved smell in the lab-kitchen! I think I will switch over to a clear variant of liquor for a more professional look :D

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#19 Post by MicroBob » Sat Aug 17, 2019 8:45 pm

Hi together,
I found some more time to look through Doron's Pleurax.mounted slide. It is always astonishing, how long one can look at a strew slide and still found new diatom forms sponge needles and other interesting oblects.

Here a fragment od a beautilful diatom, there was just this single one so far.
Zize of complete diatom: ca. 125µ

Bob
Attachments
DSC_2564bearbeitet1024.jpg
DSC_2564bearbeitet1024.jpg (150.2 KiB) Viewed 744 times

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#20 Post by MicroBob » Sat Aug 17, 2019 8:50 pm

Here a very long diatom, several hundered µ.
Attachments
Diatomee lang, ca. 400µ.jpg
Diatomee lang, ca. 400µ.jpg (151.43 KiB) Viewed 743 times

User avatar
75RR
Posts: 6063
Joined: Sun Oct 12, 2014 2:34 am
Location: Estepona

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#21 Post by 75RR » Sat Aug 17, 2019 10:33 pm

Again some very nice detail. Are you rotating the stage to look for more a favorable orientation of the diatoms?

There is some fine detail on the long diatom just on the edge of visibility.
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#22 Post by MicroBob » Sun Aug 18, 2019 9:27 am

I took these photos with my Nikon 1J5 that is directly connected to an older Leitz Periplan objective and 8mm spacer to come to a 10mm eyepiece pickup point. I just plunged the camera in the bino tube an can rotate it if necessary to get the object in the frame. Right now I have mounted a rectangular stage with low controls that doesn't rotate, so I couldn't rotate the stage to look for a better alignment of striae and DIC orientation.

I think with COL there would be a bit more detail visible and also the combination of DIC and oblique gave a slight improvement.
In this case I wasn't after maximus resolution but was toying around with the Zeiss DIC "alt" and different objectives.
If I remember right the diatom fragment was taken with a Leitz NPL Fluotar 100:1 1,32 and the needle shaped, very long diatom with a Zeiss West 40:1 0,85 oil.
DIC offers good contrast and resolution together with a very pleasing look of the images and is fairly easy to set up. But I don't have the impression that it offers new records in diatom resolving power.

Päule Heck, who makes remarkably good diatom images has a Zeiss DIC "alt" too. I asked him how well he gets on with it and his reply was that he doesn't use it a lot. For him bright field with a little bit of oblique is the preferred technique for diatoms.

Bob

User avatar
75RR
Posts: 6063
Joined: Sun Oct 12, 2014 2:34 am
Location: Estepona

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#23 Post by 75RR » Sun Aug 18, 2019 9:51 am

Found some very nice images by Päule Heck and others:

http://www.mikroskopie-bonn.de/themenga ... html#a4736

I believe oblique can also benefit from a rotating stage.
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

MichaelG.
Posts: 1397
Joined: Mon Apr 10, 2017 8:24 am
Location: NorthWest England

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#24 Post by MichaelG. » Sun Aug 18, 2019 12:45 pm

75RR wrote:There is some fine detail on the long diatom just on the edge of visibility.
Here's a nice word to drop into conversation: "limen"

Not commonly used, but singularly appropriate to the threshold situation that you describe.

Strangely enough, most people seem to be familiar with the word 'subliminal' ... but would struggle to define 'limen'

[/digression] MichaelG.
Too many 'projects'

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#25 Post by MicroBob » Sun Aug 18, 2019 1:35 pm

Hi together,
I tried to make a comparison to get a better understanding about how much Zeiss DIC "alt" can manage in fine detail compared to other illumination methods. The objective was a Leitz NPL Fluotar 100:1 n.a, 1,32 which works, but with some colour gradient. Each stacked from dozens of images.
First image was taken with oblique lighting, an objective can lid was leaving a 3/4 moon free by partly blocking the light exit at the field aperture.
Second image with DIC.
In both cases I used the green channel, and did sililar editing to the image.

Which do you prefer?

Bob
Attachments
pysharp_sup0_al1Diatomee 1 schief beleuchtet 100er 1_32-RGB.jpg
pysharp_sup0_al1Diatomee 1 schief beleuchtet 100er 1_32-RGB.jpg (106.3 KiB) Viewed 675 times
Diatomee 1 DIC 100er 1_32-RGB 1024.jpg
Diatomee 1 DIC 100er 1_32-RGB 1024.jpg (135.25 KiB) Viewed 675 times

User avatar
75RR
Posts: 6063
Joined: Sun Oct 12, 2014 2:34 am
Location: Estepona

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#26 Post by 75RR » Sun Aug 18, 2019 1:55 pm

Both images have areas where one seems a little better than the other but neither one (this no doubt due to the diatom chosen for the test) has the fine detail needed for a valid comparison.

I like the idea of a comparison however. If we could all use the same diatom, say from a Kemp 8 Test Slide then all our results would be comparable.

We could compare Oblique, 2 versions of DIC and maybe COL

Perhaps a new thread would be indicated.
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#27 Post by MicroBob » Sun Aug 18, 2019 6:33 pm

My test was more oriented towards the task at hand: To get a nice image of diatoms like this and identify the most suitable method.
For me the result is that my DIC set is usable for diatoms but not necessary. This is about what I did expect and I chose diatoms as a first subject with the DIC set as they stay in place when mounted in Pleurax and the like. :D
I have moved away from too much testing as this easily lead on a track away from the occupation with the subject itself.
The next subject I want to have a look at are desmids - I will receive a sample with some different forms on tuesday.

User avatar
75RR
Posts: 6063
Joined: Sun Oct 12, 2014 2:34 am
Location: Estepona

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#28 Post by 75RR » Sun Aug 18, 2019 6:48 pm

I have moved away from too much testing as this easily lead on a track away from the occupation with the subject itself.
The next subject I want to have a look at are desmids - I will receive a sample with some different forms on tuesday.
Fair enough, looking forward to your desmid images.
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

Hobbyst46
Posts: 2031
Joined: Mon Aug 21, 2017 9:02 pm

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#29 Post by Hobbyst46 » Thu Aug 22, 2019 7:12 pm

MicroBob wrote:Hi together,
I found some more time to look through Doron's Pleurax.mounted slide. It is always astonishing, how long one can look at a strew slide and still found new diatom forms sponge needles and other interesting oblects.
Here a fragment od a beautilful diatom, there was just this single one so far.
Zize of complete diatom: ca. 125µ
Bob
Bob, special thanks for this image and the next one (the long narrow diatom). They are very nice. The first one - file _2564 - is fascinating, I have never seen this diatom in my slides, perhaps because contrast was too low. I only have phase, not DIC, and these photos are very nice DIC images.
I hope you do not mind, I add below a phase contrast images of the second diatom - same type of needle-like diatom (I think it was taken with a 63X1.4 Planapo oil objective), where the dots are barely visible, and not really resolved. Your DIC works better than my phase contrast in this case; although perhaps the two diatoms are at different distances from the coverslip.
Attachments
Phase contrast.jpg
Phase contrast.jpg (427.91 KiB) Viewed 582 times
Zeiss Standard GFL+Canon EOS-M10, Olympus VMZ stereo

MicroBob
Posts: 1283
Joined: Sun Dec 25, 2016 9:11 am

Re: Diatoms from the Mediterranean Sea, Israel, alternative cleaning and mounting technique

#30 Post by MicroBob » Thu Aug 22, 2019 7:46 pm

Hi Doron,
your phase contrast and this slide work very well together, very good contrast and little halo effect. Was this slide mounted in Pleurax or in NOA61?
As far as I know phase contrast limits the top resolution slightly due to the size of the phase ring. My DIC system does the same due to the limited size of the prisms in the condenser. So there is a bit more to gain with COL, but the DIC setup is quicker and more repeatable. A quick try with oblique light and with an oiled dark field condenser gave no overall better image. A faint idea of more resolution but also artefacts and a less clean view. I still use the ancient 15W bulb of the Phomi and it doesn't put out nearly enough light for high power dark field.

When I tried to resolve Amphipleura pellucida it was very important where in the strew slide the diatom was. Slightly away from the cover slip and A.p. was not resolved any more.

I like these needle shaped diatoms for the very graphical look of the striae.

Bob

Post Reply