Evaluation of two different methylmethacrylate processing, infiltration, and embedding techniques on the histological, histochemical, and immunohistochemical analysis of murine bone specimens

Abstract

Specific histological, histochemical, and immunohistochemical techniques are often required to better understand bone pathology as well as normal bone growth and repair. Various tissue-processing, infiltration, and embedding methods are more suitable for one type of analysis than another. Here, we examined the differences between two separate methylmethacrylate (MMA) processing, infiltration, and embedding techniques on six types of histochemical and immunohistochemical protocols. In particular we assessed the effects of rapid (rMMA) and slow MMA (sMMA) tissue processing, infiltration, and embedding in tibiae sections routinely stained with hematoxylin and eosin, toluidine blue, and Von Kossa and found that the sMMA method was more suited for each of these stains. Next, we examined the effect of tissue processing, infiltration, and embedding on enzymatic staining and found that alkaline phosphatase activity was darker and more intense in the sMMA sections but more specific and sharper using the rMMA method. Acid phosphatase activity was visualized more readily with the rMMA method. Finally, the rMMA method allowed for improved antigen preservation for osteocalcin immunohistochemical labeling.