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OfflineZwinst
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microscoping mycelium on agar plates without opening them - possible?
    #28412249 - 07/29/23 01:54 AM (5 months, 27 days ago)

Hi, i finally want to get started creating monokaryons for crossing.
I realise i have to check the final mycelium for clamp connections and that i need at least 300-400x magnification to do it. Problem is, i would have to remove some mycelium from the plate, put it on a slide and then put it under my microscope. Which costs a lot of time and effort and creates opportunity for contamination.
With the 10x lens (125x) i can put the whole plate in, but thats not enough magnification. With the 40x lens (500x) the working distance is too short to get anything in focus.

Are there affordable microscopes or lenses that enable me to just use the whole plate?


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OfflineVP123
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Re: microscoping mycelium on agar plates without opening them - possible? [Re: Zwinst] * 1
    #28412360 - 07/29/23 05:00 AM (5 months, 27 days ago)

There are long working distance objectives. I have never used them but they exist. They are not cheap as far as I know.

I  assume you are using an upright microscope rather than an inverted microscope. You could try flipping the petri dish in the microscope stage and see if you can get a clear enough image through the agar layer. Depending on your agar it may be enough for your purposes. Or you may have to make thinner and more transparent agar layers. I just hope this works for you.


Edited by VP123 (07/29/23 05:01 AM)


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OfflineZwinst
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Re: microscoping mycelium on agar plates without opening them - possible? [Re: VP123]
    #28412438 - 07/29/23 06:42 AM (5 months, 27 days ago)

I already tried flipping them around. The distance is still way too big. Pouring thinner isn“t going to help either, i have to get the hyphae within less than a milimeter in order to focus on them.
Yes, i have an old standard upright microscope (with 70s leica lenses and newer zeiss eyepieces) and i know of those expensive long working distance objectives. The cheapest ones i found were still 400 bucks though... thats not happening anytime soon.

I was just wondering if anyone has found a workaround yet or something that is affordable.

I have also seen those cheapo chinese ccd microscopes with lcd screens. They claim between 400x and 600x depending on the model and a working distance of >15mm, which would be perfect. Has anyone experience with those? If they are good enough to see the clamp connections, i would buy one. But somehow i doubt that very much.


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Offlinetrippleblack
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Re: microscoping mycelium on agar plates without opening them - possible? [Re: Zwinst]
    #28413187 - 07/29/23 08:02 PM (5 months, 27 days ago)

I purchased long working distance lenses to properly work under my am scope. they where moderately expensive but well worth the money.

i have never had a big problem flipping thin plates and analyzing 400x, you see a mono mark that spot with a marker on the petri, extract that agar section, and hope that it remains a mono. Thin pours are best.

My favorite method is just removing the cover off the petri dish and lowering the microscope all while in front of the flow hood -this has never led to infection -i would just wipe the scope down, and place it in front of flow hood for a couple hours before use.


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Invisiblecovertjoy

Registered: 07/09/23
Posts: 272
Re: microscoping mycelium on agar plates without opening them - possible? [Re: Zwinst] * 1
    #28413360 - 07/29/23 11:04 PM (5 months, 26 days ago)

I'm still learning but so far I've had the most success with preparing a slide after doing a transfer and viewing with darkfield 400x. So I do my transfer and then create the slide from what remains in the original dish after the transfer is complete. I take a small square of agar/mycelium about 1-2mm square, put it on a slide and then press a slide cover onto it to flatten it out.



Excuse the poor quality image, the camera that came with my second hand microscope leaves a lot to be desired.


--------------------


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OfflineJust M
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Registered: 08/06/23
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Re: microscoping mycelium on agar plates without opening them - possible? [Re: covertjoy] * 1
    #28438116 - 08/18/23 11:48 AM (5 months, 7 days ago)

:threadmonitor:


--------------------

APE x NSS cross I created


Cluster heaven.


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