Fast and accurate protocol for histology and immunohistochemistry reactions in temporomandibular joint of rats

Objective: Propose a standard, fast and accurate protocol for the processing of the temporomandibular joint (TMJ) of adults' rats for histology and immunohistochemistry reactions. Design: Wistar male rats were perfused with paraformaldehyde (4 %). The heads were fixed in formaldehyde 10 % solution for 48 h. After that, the heads were sectioned in a sagittal plane and fixed for plus 48 h. Decalcification was performed using 20 % formic acid for 96 h and delimitation of TMJ area was done. Detailed methodology to a standard extraction and processing of TMJ to histological sections is described. Different buffers, equipment, temperature and time were tested to optimize immunostaining. Morphological preservation and antigenicity were evaluated by hematoxylin and eosin staining and immunohistochemistry reaction. Results: The current findings demonstrated that TMJ fixed in 10 % formaldehyde and decalcified in 20 % formic acid optimized decalcification processing time with preservation of cell morphology. Antigen retrieval with citrate buffer in pressure cooker (2 min at 100 degrees C and 5 min at room temperature) demonstrated the best protocol to preservation of the structures of TMJ. Conclusions: This work demonstrates in detail a methodology of a fast and accurate TMJ processing for histology and immunohistochemistry reactions that guarantee tissue integrity and quality of staining.