Did I accidentally create phase contrast?

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Did I accidentally create phase contrast?

#1 Post by paradoxbox » Fri Jun 16, 2017 12:15 pm

Hi all,

I got my first microscope a few days ago, an old Japanese made student level light-scope. The quality is quite good, metal focus gears and the focus mechanism is -very- smooth. But it's a very basic model with the little rotary-telephone style diaphragm under the stage. I don't have much info on the objectives but I assume they are simple achromats - 4, 10 and 40x (I am considering buying a 100x oil lens soon). 10x and 15x wide field oculars.

The scope is an old one made in the 1980's so it has a mirror at the bottom and I use my iphone's LED to light up slides.

Anyway I was checking out some cheek cells at 600x, and while they were OK to look at, with OK contrast, I started messing with the mirror angle. The mirror is old and a bit dirty and when one of the shadowy parts passed over the cheek cell, the details of the cell popped out, it was a major difference. There also appeared to be a lot of chromatic aberration or halo effect around the cell.

This effect was not at all like what happens when I simply use a narrower aperture on the diaphragm - that just tends to make the picture darker. This was clearly higher contrast and it was easy to see the varying height of the middle of the cell.

What I saw resembled this wiki image very closely, though my scope has better contrast than the left side image. The right side image was nearly identical to what I was seeing.

https://en.wikipedia.org/wiki/File:Brig ... _image.jpg

Have I stumbled on to a kind of cheap hack to get phase shift?

The only thing I can think is going on is that the small grime spot on the mirror was small enough to cause some diffraction effect coming through the slide. That might explain the CA and halo effect. But I am not sure. Any ideas?

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Re: Did I accidentally create phase contrast?

#2 Post by zzffnn » Fri Jun 16, 2017 12:30 pm

Welcome to the forum!

I think what you accidentally produced was oblique illumination (light filling part of the objective aperture, causing shadow relief effects). Pull off eyepiece and peek down eye tube, you may see the effect yourself.

For phase contrast, you need light-absorbing phase ring inside (specially made) objective, matching (in size) phase annulus at condenser, then align them carefully. All 3 conditions have to be there for phase contrast to happen.

It sounds like your scope does not have a condenser. If that is true, it won't utilize the full potential of a 40x NA 0.65 objective, not to mention a 100x NA 1.25 oil objective.

I do a lot of extreme oblique illumination and found that it is difficult to avoid CA and halos in oblique light, unless highly corrected apochromatic objectives and maybe even achromatic aplanatic condenser are used.
Last edited by zzffnn on Fri Jun 16, 2017 1:16 pm, edited 1 time in total.
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Re: Did I accidentally create phase contrast?

#3 Post by 75RR » Fri Jun 16, 2017 1:03 pm

Hi paradoxbox, welcome.

Agree with zzffnn, a slightly misaligned mirror is a good way to discover oblique lighting.

One of the most useful and cost effective illumination techniques.

Do post photos of your microscope in the My microscope section - we all like to see them.

You might find a 20x more useful than a 100x - make sure whichever one you get is parfocal with the ones you have.

Have a look around the Resources (online, books etc.) section - there is a lot of useful information there.
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Re: Did I accidentally create phase contrast?

#4 Post by apochronaut » Tue Jun 20, 2017 11:35 am

You can make up various shaped cut outs from a sheet of opaque paper, thin cardboard or stiff plastic film : similar to what are being sold here : http://www.ebay.com/itm/Microscope-Dark ... xyY3ZRknos

With some of the cut outs, precisely arranged, you will get an enhancement of the effect you have noticed and restrict the amount of chromatic aberration that the oblique illumination is causing. If you leave a handle on an otherwise circular mask for your cut outs, you can use your disc diaphragm as a carrier( largest aperture) and manipulate the cut outs over the hole to adjust the results. I would recommend off-set slits as well.
You will likely have to increase the intensity and refine your illumination, in order to get the best results.

The overall size of your custom masks, will have to be proportionate to the size of the aperture in your disc diaphragm, so smaller than the ebay offering.

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Re: Did I accidentally create phase contrast?

#5 Post by Moose » Wed Jul 26, 2017 11:26 pm

I would be near impossible to accidentally produce phase contrast as the objective has a ground ring inside and the condenser has a stop that aligns with this ring. I agree with others, you have produced a form of oblique lighting and a very effective technique at that.
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