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Immunofluorescent quantification of ribonucleotide reductase M1 subunit and correlation with DNA content by flow cytometry by Graham J. Mann; Marlen Dyne; Elizabeth A. Musgrove is a Biochemistry, Genetics and Molecular Biology article available to read on EtoBox.

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The cytoplasmic enzyme ribonucleotide reductase is essential for DNA synthesis, and its activity is strongly correlated with cellular proliferation. This paper describes a flow cytometric technique for the simultaneous measurement of DNA content and the M1 subunit of ribonucleotide reductase. Data are presented for cycling cultured human leukemic lymphoblasts in which M1 is constitutively expressed, and peripheral blood lymphocytes in which it is only detectable with certainty after mitogen stimulation. The choice of fixation procedure strongly influenced the amount of M1 subunit de-tected. Paraformaldehyde (PF) at concentrations of 2% (wlv in PBS) or greater provided optimal results. Fixation at 3'7°C was significantly more effective in preserving M1 than fixation at room temperature or 4°C. These variables are shown to have affected cytoplasmic retention during postfixation processing. Their relevance to the flow cytometric measurement of other intracellular components by this procedure are discussed. Key terms: Cell cycle, cytoplasmic proteins, paraformaldehyde, propidium iodide Ribonucleotide reductase catalyzes the conversion of ribonucleoside diphosphates to their correspondi

Who reads Immunofluorescent quantification of ribonucleotide reductase M1 subunit and correlation with DNA content by flow cytometry?

Author

Graham J. Mann; Marlen Dyne; Elizabeth A. Musgrove

Publisher

John Wiley and Sons; Wiley (John Wiley & Sons); John Wiley & Sons Inc.; John Wiley & Sons Ltd.; Wiley (ISSN 0196-4763)

Published

1987

Language

EN

Field

Biochemistry, Genetics and Molecular Biology (Life Sciences)