Studies of platelets with heavy metal impregnation techniques

Various methods of heavy metal impregnations were performed on human platelets. The optimal technique consisted of glutaraldehyde fixation, incubation in warm uranyl acetate at a pH of 3.5, followed by a double solution of lead and copper, and finally overnight immersion in cold osmium tetroxide. Semi-thin sections, viewed at 90 kV, revealed three types of platelets: (1) 'reticular' cells, with a prominent tubular network and very dark granules in a pale cytoplasm; (2) 'dark' cells, with an electron-dense cytoplasm; and (3) 'pale' cells, with microvesicles and non-staining granules. Pre-treatments with EGTA, aspirin and various platelet activators altered the appearances and proportions of the three cell types. A cell-partitioning two-phase polymer system showed that the sub-grouping is related to surface membrane properties, the cells retained in the top phase being exclusively type 2 'dark' cells. The changes in cell type distribution produced by activation show that metal impregnation may be a useful method for studying structure-function correlations in platelets.