Can someone help me with some cryosectioning issues?
I made mouse Achilles tendon tissues in vitro and my protocol is already validated. The skin is presenting some cracks, I think something is wrong with the freezing protocol, as followed:
After the tendon is ready, I've placed it in 4% PFA overnight.
Then 15% sucrose incubation until tissue sinks, after that I move the tissue into 30% sucrose and wait again until it's sinking.
After embedding on a mould with OCT and immediately store the mould in -80 overnight. Before I cut the tissue the sample has been put in -20 overnight.
From the photos I think something is wrong with this process, can someone help me?
While processing the tissue, there is possibility that your tissue was too thick, make 2mm thick tissue, that may help you specially getting 30% sucrose in.
I did frozen section on mouse tendon (a reprint enclosed), Please see Figure 3
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