[Histonet] cryoprotection before MMA embedding at -20 =?iso-8859-1?q?=B0C?=


Hi everyone
i use a protocol for MMA embedding at 20°C since a few months.
 and this protocol says to put simples in phosphate buffer 7.4 with 10% sucrose overnight at 4°C after fixation in paraformaldéhyde 4% for 3 days and before dehydratation and preimpregnation at 4°C,  emebdding is done at -20°C.
Last time i forget the step of cryoprotection with sucrose, and after cutting and staining my embedding simples, i didn't see any cells, no one cell !!! i did never see that before !!!!
Do you think that is because of no cryoprotection ?
Does a lack cryoprotection induce disparition of cells ? the tissue was very thin... 
any help would be very usefull
Thank you very much
Myriam Baali
Natural implant

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