Optimizing Images (M.orbicularis?)

[TravisH]

Good morning Macro, and Micrographers .

I had two questions, but the primary question is about technique which is why I have put it in this section of the forum, if it is not appropriate please let me know and move it if needed.

The first question, and likely the easiest is does anyone have an ID on this, to me it sort of looks like M.orbicularis but I am not 100% sure.

The second question is the far bigger technical question. This image was from memory about a 10x image on a Nikon D5300 DSLR which was focus stacked from say 10 images. These images went into Helicon Focus, and I tried Depth Map, and also Pyramid and came up with essentially the same image (below). When I look at the image, with the exception of seeing how dirty my objectives are, I can't help but be a little disappointed in the image since to me it looks almost too 'cartoonish', like it was drawn. Is that just something I need to expect from these sort of images, or is that more to do with my technique. If it is a technique related issue are there tips on how to get around that?

Many thanks!

[zzffnn]

Does the image from vusual eyepieces look the same (size and quality wise)?

If so, and your scope does not have Kohler light path, I suggest:

1) raise your condenser higher, to almost touching slide;

2) open up condenser iris more, you can open iris completely if you are stacking;

Or if you are not stacking, open condenser iris and use an oblique mask to block about 2/3 of the condenser bottom entrance; that will give you good resolution and contrast;

3) use a blue filter on top of light source, if you want white background

If your visual eyepieces produce much better image of very different image size, then your photo tube and camera setup may need improvement.

You may want to post a photo of your setup and provide more details.

Sometimes, your samples contains sand particle and slides/cover slips also may have dust particles, all if which may show up in the final image. So it may not be your objective being dirty. But you can carefully clean the lenses with breath and cotton swap.

[Charles Krebs]

So many variables to consider. It would be good to have some idea of the microscope and optics involved.

Without having more info than you have provided, the first two things I would look at would be the aperture you had set on the condenser. Closing the aperture down too far will give a more "contrasty" image but it can really destroy image resolution. What procedure do you use to determine where to set the microscope condenser diaphragm?

The second thing I wonder about is vibration caused by the camera mirror and shutter.mechanisms. What is the procedure you use to eliminate or reduce camera vibration?

[TravisH]

Thanks guys, The microscope uses Kohler illumination, however in this case I might have had the condenser too far away from the stage. The image looked fine similar through the eyepeices so I don't think it is camera specific.

I have taken images of other things using the same setup and that came out fine. I use a combination of a stable desk, but also shutter delay to try and limit the 'shake' factor.

Thanks for the suggestions, I will fire up the microscope again tonight and try again with the condenser closer to the slide. Always hard to find the right amount of contrast!

[Charles Krebs]

If your microscope does have Köhler illumination, keep in mind that the condenser height is not arbitrary, but should be set to a specific height using a simlpe proper set-up routine. (Once this is established you can vary it a little in some cases to see if anything changes for the better, but usually best to have it at the "proper" height).

The aperture size used can be quite important, if in doubt pull an eyepiece and look down the tube and use the 70-80% "rule".