ENZYME-HISTOCHEMISTRY OF RAT MAST-CELL TRYPTASE

Fixation and staining conditions for rat mast cell tryptase and its hi stochemical distribution in different rat tissues were investigated. P rostate, skin, lung, gut, stomach and salivary glands were fixed in ei ther aldehyde or Carnoy fixatives and then frozen or embedded in paraf fin wax. Preservation of tryptase enzymic activity against peptide sub strates required aldehyde fixation and frozen sectioning. Of the pepti de substrates examined, z-Ala-Ala-Lys-4-methoxy-2-naphthylamide and z- Gly-Pro-Arg-4-methoxy-2-naphthylamide proved the most effective for th e demonstration of tryptase. Double staining by enzyme cytochemistry f ollowed by immunological detection of tryptase showed that, in all try ptase-containing mast cells, the enzyme is at least in part active. Co nventional dye-binding histochemistry was used to confirm the identity of mast cells. Aldehyde-fixed mucosal mast cells required a much shor ter staining time with Toluidine Blue if tissue sections were washed d irectly in t-butyl alcohol. Double staining by enzyme cytochemistry an d dye binding showed that tryptase is absent from mucosal and subepide rmal mast cells, which are also smaller in size and appear to contain fewer granules than connective tissue mast cells. This study demonstra tes that rat mast cell tryptase, unlike tryptases in other species, is a soluble enzyme. It is stored in an active form and is absent from s ome mast cell subpopulations in mucosa, skin and lung. (C) 1998 Chapma n & Hall.