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Tipps Gefriermikrotom

Begonnen von mlippert, Mai 14, 2021, 22:56:42 NACHMITTAGS

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mlippert

Hallo,

Heute ist diese weiße Schönheit bei uns zu Hause aufgekreutzt. Zwar nehme ich das gleiche Modell auch im Institut, aber ich habe durch YouTube feststellen müssen, dass ich so viel nicht weiß, über Gefrierschnitte und Cryostaten. Also wenn ihr gute Tipps für diese Geräte habt, immer her damit! Ein netter Forenteilnehmer hat mir auch schon eine Einladung zu einem Leica Seminar geschickt.
Die hauptsächliche Funktion des Geräts sind schnelle Schnitte von Tumorgewebe in der Tierarztpraxis, aber vielleicht darf ich ja nachts auch Mal was Privates schneiden, wenn das Weibchen nicht hinschaut. 😁

Rene

Nice! For best results, use fixed material, eg with formalin and infitrate overnight with 15-20% sucrose as a cryoprotectant. Then freeze on a stub and start sectioning. There should be a plastic plate above the knife which prevent rolling up of the sections. Try picking up on a dry slide, or on a slide with a drop of mountant (50% glycerol +DAPI works nicely!). In my hands with plant material (which easily shatters), dropping in mountant gave less damage than picking up on a dry slide, before mounting. But it is probably less problematic with animal tissue. I didn't get much thinner than about 15 um at around -15oC.

Good luck,
René

mlippert

#2
Oh, plant material is extra interesting to me as I never managed with my primitive hand held microtomes to slice as thin as I wanted. I'll try the glycerol idea.
Do you know how to adjust the anti roll plate? What's too far back? Too far out has the glass scraping the object, but where is the best spot? If you have one made from plastic, throw it out and replace with a piece of conductive ITO glass. That keeps the sections from sticking to it. Commercial plates are all conductive. Worst case, use an old dichroic IR filter.

Should I actually power it down if it's not being used for a few days? Judging from the loudness, it's quite power hungry...
Do I still have to dry the microtome if I let it warm up with the lid closed?

Best,
Michael

Ps: attached is the first cut through some skin, left side is normal, right side has some sort of lesion, I forget which...

Rene

Hope it's not a sample from your finger ;-)

Never had a problem with the roll plate (perspex), adjusting was not much more than just out of the way of the surface of the block, and slightly above the knife (mm or so), that the section slides through without a problem. Adjustment  'on the go'.

I suppose let it come up to roomtemperature for a few days and then open the window to dry up completely.
I never needed maintaining it myself, but you'll need some thinner greases to be able to work it at low temp.

Good luck,
René

rlu

#4
Zitat von: Rene in Mai 14, 2021, 23:44:59 NACHMITTAGS
Nice! For best results, use fixed material, eg with formalin and infitrate overnight with 15-20% sucrose as a cryoprotectant.
Hi Rene,

sucrose is a mixture of fructose and glucose. Glucose is forming crystals.
I don't know if this is an issue in your case.

Rudolf

mlippert

Ich dachte sucrose ist einfach Haushaltszucker? Jedenfalls nehme ich den seit Jahren für meine Gehirne. Nie Probleme gehabt...