The specificities of antibodies raised in rabbits against rat mast cell proteinase 1 (RMCP 1) from connective tissue mast cells (CTMC), and against RMCP II from mucosal mast cells (MMC), were analysed by SDS-PAGE and Western blotting. Significant cross-reactivity was detected, and was eliminated by affinity purification and cross-absorption techniques. The resultant F(ab')2 antibodies, monospecific for each enzyme, were used for the immunohistochemical localization of RMCP I and II in rat tissues. Cells in skin, tongue, intestinal serosa and lung parenchyma which, by histochemical techniques, have been identified as CTMC, contained RMCP I exclusively. Cells in jejunal lamina propria and bronchial epithelium, previously classified as MMC, contained RMCP II. The results demonstrate the feasibility of distinguishing mast cell subsets by their content of serine proteinases.