MicrobeHunter.com Microscopy Forum

The microscope used here is a modified Amscope T400. The difference between this scope and the T490 is that it is non simul-focal. One must switch from eyepiece to camera and both can not be viewed at the same time. The other difference is the price. The 400 doesn't cost as much as the 490. Both are finite scopes.

Because you folks were nice enough to tell me what I had to do, I modified the scope from 20 watt halogen to 3 watt LED. It is can be made to be extremely bright. That extra iintensity comes in handy for phase, dark, and oblique lighting.

I changed all the objectives to PLAN which didn't seem to make much difference until I added PLAN eyepieces. My, what a difference!

Specimen images are taken using a 100x PLAN, IRIS objective. Condenser iris is set to about 50%. One image is with olive oil ( R.I. =1.46 ) and the other image is dry. There is not much difference between them that I can see with my old worn out eyes.

Results.
1. Modification of light source - worthwhile.
2. Use of plan imaging - worthwhile
3. Use of Iris Objective - worhtwhile

A good morning opver all. Have fun, Greg

Hi Greg,
as you imagine we now all want an image made with proper immersion oil.
In my eyes the image with the oilve oil is considerably better than the image with the objective used dry.

Bob

I'm pretty surprised just how close the dry image is to the immersion. Incidentally the globular-looking red-stained substance is I think a globular-form of tannin, very often seen in my slides too, in varying stages of formation and deposition within plant cells. Protective (against herbivory I think) for the plant, although perhaps storage/waste?

Very nice 'scope and images.

Greg - I gather this is an Amscope 100x oil objective with a built-in iris? If so, do you recall the setting of the internal iris? It would be useful to know, between the condenser iris and any internal iris about what numerical aperture ended up being used.

It says Plan 100/1.25 oil-0.5
160/0.17
I set the iris at about 50%.
I didn't use regular immersion oil because I use it only for strict research. A pint of Olive oil costs me six bucks. A tiny bottle of immersion oil is 20.

Aha - that may be why the contrast of the dry image is good.

You can pick up a nice zeiss 100x oil with iris for around $30, and a Neofluar version for not much more.... as long as it RMS you can run anything on it, and I doubt the Amscope eyepieces have any correction in them as they buy from jobbers...

In closing both your objective iris diaphragm and your condenser iris diaphragm, you are reducing the N.A. of the objective. I don't know actually how much in this case but if you say the diaphragms are at about 50% , then your objective N.A. is no longer 1.25, it is about .85 roughly. The condenser about the same as well. Therefore, your objective's potential resolution is considerably reduced from what it's capability is using actual immersion oil, with the iris wide open and the condenser oiled to the slide, with it's diaphragm wide open. Immersion mediums of any kind , provide the capacity to achieve an N.A. greater than 1. In limiting the objective N.A. by closing off the diaphragms to below 1, no immersion medium is necessary to achieve the maximum N.A. of the objective, thus both dry and oiled the objective performance will seem similar, yet poor. There is still some advantage of the olive oil, in that it helps to reduce spherical aberration somewhat, thus the image taken with the olive oil is slightly better than the dry image. Closing the diaphragms that much also increases contrast , so although the resolution is poor, there is a kind of illusion of there being a tidy microscope image.
If you open both diaphragms ,oil the condenser to the slide with immersion oil and then take a picture with a dry 100X (1.25) objective , then do the same with the objective immersion oil immersed as well, you will see a dramatic difference between the resolution of those two images as well as between the fully immersed image and the previously posted images. That comparison shows the true relationship between an immersion objective immersed and an immersion objective dry.

F.y.i. the maximum possible resolution for 500 nanometer light for each of the 4 scenarios above assuming the objective and conthedenser used in the first two photos were working at .85. N.A.
. 1) .85 objective, .85 condenser. both dry------------------------------------------------------------.359 microns but confounded by spherical aberration so somewhat less
2) 1.1 objective immersed in olive oil, .85 condenser dry.-----------------------------------.313 microns but also confounded by s.a.,
3) .95 objective unimmersed , 1.25 condenser oiled to slide.----------------------------.316 microns but confounded by s.a. and flare from the wide open condenser. There is no benefit to oiling the condenser and not the objective, since the full N.A. of the condenser cannot be achieved if the objective is incapable if utilizing it.
4) 1.25 objective oiled to cover slip and 1.25 condenser oiled to slide.---------------.244 microns

When saving on oil. Olive oil has some colour and has one of the lower n's of the culinary oils plus it is one of the most expensive. Peanut and corn have a little higher n's and are almost water clear.

Thank you very very much! I understand it now.

Hey Greg, bringing up this old post because you mentioned something interesting...

I have a T490 and you mentioned PLAN eyepieces for your scope that made a large difference. My scope accepts 23mm mounting diameter eyepieces, and I am having difficulty finding these PLAN eyepieces you talk about... I am only finding PLAN ones for larger diameter eyepieces. Do you have a source for them?

Josh