Detection of cytoplasmic and surface membrane markers in cells of some human hematopoietic cell lines.
The cells of some human leukemia-lymphoma T cell lines (JURKAT, MOLT4), B cell lines (DAUDI, U-266) and of myeloid U-937 cell line were characterized for their surface membrane and cytoplasmic marker profiles. The usefulness of some fixation and permeabilization methods of cell membrane for detection of cytoplasmic markers by flow cytometry was studied. The methods of cell fixation in suspension were found to be more sensitive than the methods of cell fixation in smears. With the very short buffered formaldehyde-acetone (BFA) fixation used in this study an optimal penetration of the monoclonal antibodies (MoAbs) through the plasma membrane and specific binding to the appropriate structures were achieved. CD22 antigen was detected in cytoplasm but not on membrane of DAUDI cells. In another B cell line, U-266, CD22 antigen was present both in cell membrane and cytoplasm. The marker corresponding to anti-CD19 MoAb was detected in cytoplasm but was absent on membrane of U-266 cells. Furthermore, the antigen estimated by anti-CD3 MoAb could be detected intracellularly in cells of both T cell lines tested, while it was absent on cell membrane of these cells. The phenotypic study of U-937 cells showed that the majority of cells expressed myeloid associated antigens. In our study the CD14 marker detected on cell surface membrane of U-937 cells was missing in their cytoplasm. The surface antigens remained intact after BFA fixation enabling a simultaneous detection of membrane and cytoplasmic markers in double immunofluorescence studies. Through this combination of markers minor cell populations could be detected. Human hematopoietic cell lines could serve as a reliable model system for a rapid and quantitative immunodiagnosis.
Koníková
E
E
Cancer Research Institute, Slovak Academy of Sciences, Bratislava.
Babusíková
O
O
Kusenda
J
J
Glasová
M
M
D000951:Antigens, Neoplasm / Q000032:analysis*; D000954:Antigens, Surface / Q000032:analysis*; D014408:Biomarkers, Tumor / Q000032:analysis*; D003593:Cytoplasm / Q000276:immunology; D006412:Hematopoietic Stem Cells / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D015448:Leukemia, B-Cell / Q000276:immunology*; D007951:Leukemia, Myeloid / Q000276:immunology*; D015458:Leukemia, T-Cell / Q000276:immunology*; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D014408:Biomarkers, Tumor