Random Questions about objectives and sample preparation

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anajparsat123123
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Joined: Sat May 29, 2021 2:41 am

Random Questions about objectives and sample preparation

#1 Post by anajparsat123123 » Sun Jun 06, 2021 7:15 am

Hi everyone,

I recently bought a used motic bas300 microscope (phase contrast) thanks to everyone's recommendations and I have a few questions about it and microscopy in general. This is the link to the microscope I bought: https://www.ebay.com/itm/392714315029.

1-Are the phase contrast microscope objectives removable? If so, which brightfield objective is compatible with this microscope?

2-If I could only get one brightfield objective what should I get. After that, what would be the second and third most important or best objectives to get?

3-Does the working distance on a brightfield objective need to match the phase contrast working distance if I want to interchange them (if everything else is the same)? For example, mine is a CCIS Plan Achromat Phase Contrast (inf-0.7, aperture: 0.25, working distance: 4.3) and I want to interchange it with this: https://moticmicroscopes.com/products/p ... 1001703271.

4-Should I do some preparation to raw tissue samples besides embedding? Since this is phase contrast, should I still stain the samples and with what?

5-Also, is paraffin wax used to just embed the samples so you can cut via a microtome or does it also go into the sample? If it goes into the sample does that mean I need to dehydrate it?

6-How do I make a large sample clear?

Thanks!

BramHuntingNematodes
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Re: Random Questions about objectives and sample preparation

#2 Post by BramHuntingNematodes » Sun Jun 06, 2021 3:08 pm

Embedding medium should permeate the specimen usually this entails dehydration in alcohol then soaking in wax compatible fluid such as xylene or limonene.
1942 Bausch and Lomb Series T Dynoptic, Custom Illumination

farnsy
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Re: Random Questions about objectives and sample preparation

#3 Post by farnsy » Mon Jun 07, 2021 2:40 am

anajparsat123123 wrote:
Sun Jun 06, 2021 7:15 am
1-Are the phase contrast microscope objectives removable? If so, which brightfield objective is compatible with this microscope?
Phase objectives will screw right out and you can replace them with regular objectives. The most compatible objectives will be those sold by Motic. However, there is very good chance these objectives are compatible with other Chinese-made infinity objectives, which are intended to be pretty compatible with Olympus' standard (180mm tube, 45mm parfocal, RMS threads). I have not tested the Motic with other Chinese infinity optics...it's just my suspicion.
2-If I could only get one brightfield objective what should I get. After that, what would be the second and third most important or best objectives to get?
Your phase objectives can do bright field, it just won't be quite as good. So which magnification are you using the most and wishing was a little better?

I see your microscope doesn't have a 20X objective. This is an incredibly useful objective as the magnification is just right for many things we look at, such as pond eukaryotes, but the NA is low enough that it's easy to use and things like brightfield and oblique illumination work without using a special condenser. I suspect your phase condenser has a darkfield mode, which would work with 20X. So that's my first suggestion. It kind of looks like you have an empty spot right now. That's what I'd put there.

After that it's a question of replacing one of the magnifications with something you like better (for brightfield). Are you using your 100X oil? If not, perhaps consider a 60X for that spot, or a 100X water immersion.
3-Does the working distance on a brightfield objective need to match the phase contrast working distance if I want to interchange them (if everything else is the same)? For example, mine is a CCIS Plan Achromat Phase Contrast (inf-0.7, aperture: 0.25, working distance: 4.3) and I want to interchange it with this: https://moticmicroscopes.com/products/p ... 1001703271.
Working distance is measured from the front end, so it doesn't have to match. Since these are from the same manufacturer, they will probably be approximately parfocal, which is what matters. You could always ask Motic, if you want an authoritative answer.

apochronaut
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Re: Random Questions about objectives and sample preparation

#4 Post by apochronaut » Mon Jun 07, 2021 12:44 pm

The BF objective that you linked to has a specific cover glass spec. of 1.1, since it is for an inverted microscope. You should choose one for an upright microscope that either is a broader cover medium range or one that has the same cover slip correction as the other objectives. It should be marked either oo/- or oo/.17

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