Midge larva, "polarized" muscles

Charles Krebs · Post by **Charles Krebs** » Sat Sep 26, 2009 11:46 pm

Came across this midge larva in my lake sample. The upper image shows an overall shot for reference. While examining it, the musculature was very apparent, so I decided to try a few photographs under crossed polarizers. This really provided an excellent view of these features.

The head was also interesting, especially the rather formidable looking mouth. So I photographed that as well. (Looking at ventral side.)

Olympus BHS. Top image stitched, taken with 4X. Second image taken with 10X. Last two images taken with 20X. Canon 50D, Olympus NFK 1.67X photo-eyepiece. Electronic flash.

NikonUser · Post by **NikonUser** » Sun Sep 27, 2009 1:34 am

Fantastic images of the muscles. Am I seeing the individual sarcomeres between the Z-bands in the muscle fibres? Images takes me back to high school zoology class.

The head capsule is interesting also. I am assumining this is a Chironomid Midge. Even if it isn't, the shape of the large concave indentation at the back of the capsule and the pattern/shape/number of the hypostoma teeth (the row of black mountains at the rear end of the mouthparts) is characteristic of at least the genus and may even be characteristic for the species. Lots of variation between genera amd many genera.
Blackfly (Simuliidae) larvae have similar teeth and I have always planned to compare images of these teeth from my local fauna- perhaps next year.

Franz Neidl · Post by **Franz Neidl** » Sun Sep 27, 2009 1:47 am

Hallo Charles!

I like very much the second and the fourth picture. Wonderful!
Is the last picture a stack? Was the camera directly fixed on the microscope?

Franz

Planapo · Post by **Planapo** » Sun Sep 27, 2009 1:59 am

My goodness!
Gorgeous! Especially the head is unbelievably rich in details. Looks like you'd x-rayed the bug!
Did you take these with extra strain free objectives?

Your "bad influence" again: I now think I need POL! Whoa, Betty!

--Betty

Charles Krebs · Post by **Charles Krebs** » Sun Sep 27, 2009 8:43 am

Is the last picture a stack? Was the camera directly fixed on the microscope?

Did you take these with extra strain free objectives?

The last picture is a 12 image stack. This is with my BHS which I still have placed on a large copy-stand baseboard and the camera mounted above (not hard attached) the trinocular tube. With the camera I am now using (Canon 50D) I don't think this is necessary any more, and may change this configuration.

These were taken with the S Plan Apos. According to the "rules" I should be using objectives designated for DIC, or marked for POL, (with the Olympus objective series I use that would be the S Plan Achromats or D Plan Apos). But I am very satisfied with the results I get. What is "lost", if anything, is for me, more than made up for in the other image qualities of the Plan Apos. (But I'm basing this opinion primarily on the testing I did when setting up DIC. I really should do some comparisons with more basic polarization techniques). I would also suspect that with an objective not designated or marked for DIC or POL, there could be more variability (when used for DIC/POL) between different objective samples.

Am I seeing the individual sarcomeres between the Z-bands in the muscle fibres?

Not sure what the structure is that is causing the pattern. Would not expect to see individual sarcomeres.

Charles Krebs · Post by **Charles Krebs** » Sun Sep 27, 2009 9:17 am

I'll add one more image with interesting details. (The two small orange/red circles are Trachelomonas that were attracted to the light of the microscope, and have nothing to do with the larva. Some are visible in the third image above as well).

lauriek · Post by **lauriek** » Sun Sep 27, 2009 9:29 am

Wow!

Is 'Crossed Polarizers' when you have a polarizing filter on your lights and a polarizing filter on your lens and you adjust them to be 'opposite' polarized somehow for these effects?

With this kind of setup does the polarizer have to go on top of the condenser? (Assuming you're using a condenser!)

Charles Krebs · Post by **Charles Krebs** » Sun Sep 27, 2009 9:43 am

Laurie,

Essentially you need one "linear" polarizer on either side of the subject. Typically one is under the condenser (not enough room between condenser top and slide in most cases). If there is no "holder you can just lay it on top of the light port in the base. Unless you have a specific "slot" (unusual on a basic microscope) it's a little trickier to place the upper polarizer. It needs to be somewhere before the camera sensor, and hopefully before the eyepieces so you can see the results through the eyepieces. (If you have "live view" it should be possible to judge the results on the screen if you can't find a place to put it where the effect is seen through the eyepieces). Sometimes if you remove the trinocular head it is possible to place a small glass polarizer (or a "film" type) under the trinocular head. (Remember that this is in the image path so some of the thin plastic polarization material often used for lights is not ideal for this).

rjlittlefield · Post by **rjlittlefield** » Sun Sep 27, 2009 11:14 am

Charlie, please correct me if I'm wrong about this, but I believe it works also to use circular polarizers, as long as the lower polarizer is "reversed" so that both normal outer faces point toward the subject.

Also it should work to use a linear polarizer on the bottom and a circular polarizer on the top, which would allow using inexpensive film on the bottom.

I don't have circulars to check this with, but theory has it that a circular polarizer is just a linear polarizer on the normal outer face, combined with a quarter-wave plate glued on the back face. By using two units "face to face" with the subject between them, the linear components should do the same thing linear polarizers would, while the quarter-wave plates do nothing visible.

If you can confirm or correct, that would be great.

--Rik

Charles Krebs · Post by **Charles Krebs** » Sun Sep 27, 2009 2:01 pm

Rik,

Yes that's correct. I've never actually tried, but the "subject facing" side of a circular polarizer is the linear polarizing side. As long as you have linear polarizing material directly facing the subject on both sides it should work.
(And if you rely on a camera meter reading a circular used in that orientation on top may make metering more accurate.)

Before I got the "official" microscope polarizers I used a variety of filters/films. They are not all equal when it comes to degree of extinction and neutrality of color. Even an excellent quality B+W did not provide as great an "extinction" as the Olympus ones. (Anecdotally, I have heard of some fairly inexpensive camera polarizers that supposedly worked very well.)

Never used them for polarizers myself, but Edmund has some possibly useful products in this regard:

http://www.edmundoptics.com/onlinecatal ... uctID=1912

http://www.edmundoptics.com/onlinecatal ... uctID=1558

Also, for photography I rarely fully "cross" them. If you do, you can lose the outline of the subject into the background (it all goes "black") and the "context" of the image is lost. If you are tight enough so that only the "illuminated" tissue fills the camera frame this is not an issue.

piotr · Post by **piotr** » Sun Sep 27, 2009 4:39 pm

Charlie, these are amazing pictures!

Rik, you are right - I am using Tiffen 52mm circular polarizer and it works very well both for polarized light and for DIC. It nicely fits illuminator port of Olympus BH2 stand. The quarter-wave plate is on a bottom of the filter, the linear polarizer part is on the top. As for analyzer, I am using round 23mm linear polarizer that fits a nosepiece aperture. I wouldn't say that the extinction is 100% (there is a faint blueish background for full-crossed configuration), but it is satisfactory for most of the work.

Aynia · Post by **Aynia** » Sun Oct 04, 2009 9:42 am

I'm playing catch up looking at this site!

I really love the cross polarisation of the specimens.

discomorphella · Post by **discomorphella** » Mon Oct 05, 2009 4:15 pm

Hi Charlie--

Spectacular shots (as usual...). Everyone may be interested to know that Edmund sells a decent polarizing sheet that you can cut (its VERY tough, but a sharp knife and some careful setup, e.g., securely attaching the film to a cutting board give good results) to shape.
Catalog online is:

http://www.edmundoptics.com/onlinecatal ... uctID=1912

Price is pretty reasonable too. Crossed extinction is not as good as really good grade mounted polarizers, but you can cut it out for making custom shapes to put in your condenser tray etc., so its worth checking out.

FWIW--

--David

Ooops, just noticed you'd already mentioned it, I have used the 1912 sheet for making custom polarizers for my orthoplan, and for making simpler polarizers for smaller scopes, it's worth having some around the workshop....