Table 3.

TRAP staining procedure

TRAP staining
Reagents: Various kits for TRAP staining are commercially available, like leucocyte acid phosphatase kit from Sigma Diagnostics Cat. No. 387-A Preparation of tartrate solution: mix the following components: 0.25 mL Naphthol AS-BI phosphoric acid, 1.0 mL acetate solution, 0.5 mL tartrate solution add 45 mL distilled water
Staining procedure	Time
Prepare tartrate solution always freshly according to manufacturer instructions
Incubate tartrate solution on specimen at 37°C in a water bath protected from light	1 hour
Preparation of substrate solution: mix following components 0.25 mL fast garnet GBC base solution 0.25 mL sodium nitrite solution	rest for 2 min at RT
Add substrate solution to preincubated slides and develop at 37°C	2 min
Rinse with distilled water	15 s
Counterstain nuclei with Meyer’s haematoxylin (see H&E staining steps 1–3)

RA, rheumatoid arthritis; TRAP, tartrate-resistant acid phosphatase.