Macrophage-Galactosidase and Demonstration of its Activity by Electron Microscopy
FORT DETRICK FREDERICK MD
Pagination or Media Count:
Hydrolases of alveolar macrophages are considered to play a role in early host resistance to airborne infection. This study biochemically examined certain properties of macrophage galactosidase and developed methods for the initial demonstration of its activity by electron microscopy. Cells were harvested from the lungs of BCG-treated rabbits and then evaluated for activity against conventional biochemical substrates and the histochemical substrate, 5- bromo-4-chloroindol-3-yl-beta-D-galactopyranoside. Hydrolysis of the latter results in the formation of indigo. Enzyme partition studies of cell lysates did not reveal preferential hydrolysis, pH optima, analogue inhibition, or heat inactivation curves unique to the particulate or soluble fraction. Evaluation of processing methods for electron microscopy revealed moderate solubility of indigo in propylene oxide. Fixation of single cells in buffered glutaraldehyde provided the best compromise for preservation of cell detail and biochemical activity. The techniques developed have been used for ultrastructural demonstration of galactosidase activity in alveolar macrophages and other inflammatory cells.