Immune thrombocytopenias are usually associated with normal or increased numbers of megakaryocytes in the marrow. Therefore, the mechanism(s) responsible for the destruction of circulating platelets may not affect megakaryocytes in the same way. One of the possibilities which could account for the differential effect on the cells would be the development of antibodies to components of platelet membranes which are not exposed on the surface of all megakaryocytes. To investigate this possibility, a rabbit antiserum specific for mouse platelets was tested against fresh and cultured mouse megakaryocytes by indirect immunofluorescence. This antiserum cross-reacted with 46% of fresh murine megakaryocytes and 54% of cultured megakaryocytes. Phase-contrast microscopy revealed the reacting megakaryocytes to be fully granulated with irregular contours and in the process of releasing platelets. Nonreactive megakaryocytes demonstrated smooth contours and lacked morphological evidence of thrombocytopoiesis. Electron microscopy showed that only in megakaryocytes (MK) with an irregular contour had the demarcation membrane system (DMS) reached continuity with the plasma membrane. Ultrastructural analysis of megakaryocytes from patients with ITP showed approximately 25% to 50% of megakaryocytes without evidence of injury, whereas 50% to 75% had extensive damage. In undamaged cells, platelet territories had not yet reached the peripheral zone. The DMS of damaged megakaryocytes opened to the exterior elaborating platelets. The observations suggested that some platelet antibodies react only with megakaryocytes which have reached the stage of thrombocytopoiesis. Relevant target antigens may not be exposed on all megakaryocytes before cytoplasmic fragmentation occurs.