Application and Evaluation of the Lead-Adenosine Triphosphatase Method in Skeletal Tissue

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Abstract
Application and evaluation of the lead-ATPase histochemical method in skeletal tissue has demonstrated an intracellular localization of enzyme activity. The skeletal tissue was demineralized for 72 hr. in cold 10% aqueous EDTA adjusted to pH 7.2. Frozen sections were cut and placed on cold albumenlzed slides, oriented, thawed, dried in a cool air stream, and fixed for 10 min. in cold (-2[degree] to -3[degree]C) 10% formalin buffered with Na-acetate and adjusted to pH 7.2. The sections were washed, treated with 10% EDTA for 20 min. at room temperature, rewashed, and incubated for an optimal period of 30 min. at 37[degree]C in the lead-ATP medium of Wachstein and Meisel. Following incubation the sections were washed, treated for 1 min. with 1% (NH4)2S, rewashed, immersed for 30 min. in 10% buffered formalin, dehydrated, cleared, and mounted. Evaluation of the substrate specificity suggests that other phosphatases associated with skeletal tissue do not complicate the ATPase reaction.