CHARCOT-LEYDEN CRYSTAL PROTEIN DISTRIBUTION IN BASOPHILS AND ITS ABSENCE IN MAST-CELLS THAT DIFFERENTIATE FROM HUMAN UMBILICAL-CORD BLOOD PRECURSOR CELLS CULTURED IN MURINE FIBROBLAST-CULTURE SUPERNATANTS OR IN RECOMBINANT HUMAN C-KIT LIGAND

Suspension cultures of human umbilical cord blood mononuclear cells su pplemented with c-kit ligand-containing additives give rise to a mixtu re of cells belonging to several lineages. Among those that differenti ate in quantity are mature basophils, immature mast cells, and neutrop hilic myelocytes. We used an ultrastructural immunogold method to dete ct the Charcot-Leyden crystal (CLC) protein, an eosinophil- and basoph il-specific protein, to study cells that were obtained at sequential t imes from 3 to 14 weeks in culture. Basophils (and eosinophils, which were present in smaller numbers) labeled for the CLC protein; mast cel ls did not. The labeled basophil subcellular sites included formed int ragranular, cytoplasmic and nuclear CLCs, cytoplasmic particle-filled and homogeneously dense granules, cytoplasm, nucleus, plasma membrane, and cytoplasmic and Golgi area vesicles. Individual basophil ultrastr uctural phenotypes similar to those associated with stimulated release and recovery reactions showed the expected variations in the gold-lab eled subcellular compartments. Macrophages also were labeled for CLC p rotein within endocytotic-lysosomal structures; neutrophilic myelocyte s did not contain CLC protein. On the basis of findings reported here, the combined ultrastructural morphology and immunogold phenotyping of cells differentiating in c-kit ligand-supplemented cultures allows ac curate lineage assignment of the developing cells.