Thrombin-induced secretion of platelet factor 4 antigen (PF4) was measured and its intracellular translocation was simultaneously followed by immunofluorescence. In permeable resting [human] platelets, speckled intracellular immunofluorescent staining for PF4 was observed. Addition of thrombin to washed platelets at 22.degree. C resulted in secretion of PF4 and formation of large (.apprx. 0.5 .mu.m) immunofluorescent masses. These masses moved to the cell periphery during secretion and were virturally absent at the conclusion of secretion. Ultrastructural examination of thrombin-treated platetlets revealed vacuoles corresponding in size, shape and time of occurrence to the large immunofluorescent masses of PF4. These vacuoles contained PF4 by immunoferritin staining of frozen thin sections; they therefore appear to represent the ultrastructural counterpart of the large PF4 masses. When intact cells were stained for PF4 after thrombin addition, only 5.6% of the large masses stained. During secretion, PF4 antigen is consolidated into large closed pools that appear as vacuoles in the EM.