Discrepancies between flow cytometric analysis and [3H]thymidine incorporation in stimulated lymphocytes |
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Authors: | Mária Sasvári-Székely Gábor Szabó Mária Staub Tatjana Spasokukotskaja Ferenc Antoni |
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Affiliation: | 1. 1st. Institute of Biochemistry, Semmelweis University Medical School, Budapest Hungary;2. Department of Biophysics, Medical University School of Debrecen, Debrecen Hungary |
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Abstract: | The DNA synthesis system of freshly isolated tonsillar lymphocytes and those stimulated by phytohaemagglutinin were compared by different methods. Both cell populations had high DNA polymerase α and thymidine kinase activities, as well as a high rate of incorporation of [3H]thymidine into DNA. However, the two cell populations differed when their DNA distributions were compared by flow cytometry. Freshly isolated cells contained many less (6%) cells in S phase than were found in phytohaemagglutinin-stimulated lymphocytes (18%) as detected by flow cytometry. The labelling of different subpopulations of lymphocytes was studied by sorting them electrically with a fluorescence-activated cell sorter. Analysis of the radioactivity of [3H]thymidine pulse-labelled cells, sorted according to their DNA content, showed that cells in the G1 peak of DNA distribution had a significant amount of incorporated [3H]thymidine. Sorting of cells according to their size (i.e., by light scattering) revealed that only large cells were labelled with [3H]thymidine. |
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Keywords: | Flow cytometry Thymidine incorporation Phytohemagglutinin stimulation (Lymphocyte) |
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