Hi all, This discussion of phosphate buffered formalin has raised some other questions for me. In paraffin embedding protocols for early mouse embryos, I have seen recommendations that dehydration begin with steps in 30% and 50% EtOH in 0.9% NaCl or PBS, then the 70% and higher dilutions are in dH2O.
I was told (and assumed) the purpose was to preserve osmolarity of the tissue, and minimize cell shrinkage - that this was a 'gentler' way of dehydrating. However, now it seems to me that at residual PBS precipitating in the 70% and higher dilutions would damage the tissue? What about unbuffered NaCl only? What does PBS precipitation artifact look like? Has anyone else encountered this practice, or perhaps can shed some light on the issue? Andrea Marion Graduate Student University of Illinois at Chicago --- Jennifer, the only reason I am aware of using acetate over phosphate buffering would be to minimize the precipitation that happens when phosphate buffered formalin contacts alcohol concentrations greater than 70%. It is why some automated tissue processors have a warm water flush step. The two fixes to this issue I have heard are: 1 - use 70% alcohol or lower in the step following the last fixative or 2 - use sodium acetate buffered formalin instead That's my wild guess for the day. I bet if you ask the investigator, the answer is nowhere close to this. <grin> Teri Johnson, HT(ASCP)QIHC Head, Histology and Electron Microscopy Stowers Institute for Medical Research Kansas City, MO _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet