Hello Histonetters, Currently, I am having trouble achieving good immunohistochemistry results using frozen mouse tibia sections. I use the protocol as follows:
4% PFA or 10% NBF 12-24 hrs cryoprotect in 20% sucrose until tissue sinks freeze tissue in block molds using OCT compound and isopentane/dry ice slice on cryostat (anywhere from 5-100um depending on microscopy work being completed) IHC: block (2-10% normal serum in TPBS) primary overnight PBS washes secondary PBS washes fluorescent mounting media I am using Cy3 labeled antibodies and not achieving that good staining I would like. Does anybody know a protocol or some special tricks I can use to avoid the autofluorescence? I can't seem to have any luck since slicing mouse bone tissue. I know many people have problems, but many publications have achieved good results. Thanks, Branden _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
