Microcytofluorometrical measurement of DNA content in immunologically identified micromegakaryocytes of human bone marrow
Kobayashi, Y.; Ozawa, M.; Maruo, N.; Kondo, M.
Nihon Ketsueki Gakkai Zasshi Journal of Japan Haematological Society 52(7): 1165-1169
1989
ISSN/ISBN: 0001-5806 PMID: 2482647 Document Number: 347085
We have developed a method to microfluorometrically determine the amount of DNA in immunologically identified micromegakaryocytes on bone marrow smears. Bone marrow smears were fixed with acetone-formalin buffer and immunostained with a monoclonal anti-GPIIb/IIIa antibody, followed by FITC conjugated anti-mouse IgG. After the smears were re-fixed with methanol, DAPI (4', 6-diamidino-2-phenylindole) staining was performed. Using an automatic Digital-Microfluorometer (Olympus MMSP-FR-II), megakaryocytes on the smears were identified by the GPIIb/IIIa immunofluorescence and, after changing the barrier filters, their nuclear DNA content was measured by the intensity of DAPI fluorescence, which is proportional to the amount of DNA. Using this method, we found that the DNA histogram of the megakaryocytes from a patient with myelodysplastic syndrome showed a shift to small ploidy compared with normal controls. This method may be valuable in the measurement of the megakaryocyte DNA content.