HRP Quenching Step Losing Frozen Slices - (Dec/17/2007 )
When attempting to quench endogenous HRP in our tissue slices, we lose a samples from the slides. Can anyone recommend a gentler way to do this? We currently use a product 'Froz-Pen' to coat Fisher super slides before tissue sectioning (although we have had the same problems with 'subbed' slides or poly-L-lysine coated slides). Are tissues are a little thicker (30um), and are acetone fixed for 30m following sectioning. They survive all other steps quite well. Any suggestions would be greatly appreciated.
apes (aka tespa) coating works better than poly L (and you can make large batches)
why is your tissue so thick?
It needs to be thick as we are sectioning bone to look for single or only a few fluorescent cells. Kind of a needle in a haystack approach, but it has worked so far.
sounds all too familiar
apes should work fine - didnt know you could cut bone on a cryostat - thought it would require more support than oct could provide (or do you soften it?)
the other alternative of course is take the parrafin/microtome route cos then you could bake the sections onto apes slides (v stable + good piccy's)
Yeah - bone is decalcified by soaking in in EDTA for a while. Thanks for your suggestion though.
i normally use vectabond (vecor) to pretreat my slides. i work with paraffin embedded tissue and have to use very harsh conditions like cooking slides in microwave oven and also have to quench endogenous peroxidase. it worked well.
i read the recommendations, and this product could also be used in combination with frozen sections.
maybe this is interesting for you.