Light and Electron Microscopic Demonstration of Phospholipase B Activity in the Mouse Eosinophil

Phospholipase B (EC 3.1.1.5) which hydrolyzes phospholipids in the alpha and beta positions was demonstrated in murine leukocytes using light and electron microscopic histochemical techniques. Leukocytes (neutrophils, lymphocytes, macrophages, eosinophils) were harvested from peritoneal exudates of mice. Cells were fixed in 4% calcium-formol fixative for 10 min at 4 degrees C for light microscopy and 30 min at room temperature for electron microscopy, after which they were incubated at 37 degrees C in medium at pH 6.6 containing 2 microM lysolecithin and CaCl2. The fatty acids released during the hydrolytic reaction were trapped as a calcium precipitate and were converted to a cobalt precipitate for light microscopy by treatment with cobalt acetate or to a lead precipitate for electron microscopy by treatment with lead nitrate. The reaction products were observed to be present in eosinophils and absent in neutrophils, lymphocytes and macrophages. It is concluded that the eosinophilic leukocyte is the carrier cell for phospholipase B in inflammatory reactions.