Histology FAQ

Staining, Histochemistry and Histotechnology

(Frequently Asked Questions)


Dr. John A. Kiernan
Department of Anatomy and Cell Biology
The University of Western Ontario
London, Canada



FAQ Home > Staining Methods, Histochemistry


Phosphatases in decalcified, embedded tissue


Can acid phosphatase activity still be demonstrated in formalin fixed, decalcified, paraffin embedded bone sections?

Answer 1.

Have a go, I used to stain for acid phosphatase in 1-10 æm sections of demineralized, glutaraldehyde/osmium-fixed epoxy-embedded specimens with no bother. The method was nothing special, just a standard napthol AS-BI phosphate/diazotised pararosaniline technique.

While we're at it, how about alkaline phosphatase in ethanol-fixed, methacrylate embedded sections? Try: McGadey,J. 1970. Histochemie 23. 180-184. Tetrazolium method for non-specific alkaline phosphatase. This is an excellent method; it has never let me down in any application.

Ian Montgomery

Answer 2.

I routinely do acid phosphatase staining on formic acid-decalcified GMA-embedded bones. Alkaline phosphatase can also be demonstrated in the GMA and is retained by the alcohol fixation. The problem that I have found with trying to both from the same block is that the acid phosphatase stains much better with formalin fixation and the alkaline phosphatase stains better with alcohol fixation.

I have had good results with acid phosphatase using formic acid decalcification and paraffin embedding of rodent skull. An excellent article is C. Liu et al. "Simultaneous demonstration bone alkaline and acid phosphatase activities in plastic embedded sections and differential inhibition of the activities." Histochemistry 86:559-565, 1987.

Martha Strachan
Skeletech, Inc., Kirkland, WA