Bivariate flow cytometric analysis of DNA content versus immunopositivity for ribonucleotide reductase M1 subunit in the cell cycle.
Ribonucleotide reductase (RR) is a cytoplasmatic enzyme catalyzing the reduction of all four ribonucleotides to their corresponding deoxyribonucleotides. Its activity strongly correlates to the rate of DNA synthesis. By using a specific monoclonal antibody against the large M1 subunit of RR, we assessed the expression of M1-RR versus DNA content by dual-parameter flow cytometry. The aim of this paper was to compare the variations in the immunopositivity for M1-RR during the cell cycle to the positivity for other cell cycle markers identifying either proliferating cells (Ki-67 and PCNA) or quiescent cells (statin). To do this, normal human embryonic fibroblasts in different growth conditions as well as several other mammalian cell lines (rat C6 glioma cells; mouse 3T3 fibroblasts and B16 melanoma cells; human epithelial EUE cells and mammary carcinoma MCF-7 cells) were used. The expression of M1-RR antigen was found to correlate positively with the expression of Ki-67 and PCNA, and negatively with the expression of statin. During early G1 phase, M1-RR becomes detectable by specific antibodies relatively later compared to PCNA and Ki-67; therefore, the lack of immunopositivity for M1-RR cannot be taken as an absolute indication of cell quiescence in G0.
Mangiarotti
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R
Dipartimento di Biologia Animale and Centro di Studio per l'Istochimica del CNR, Pavia, Italy.
Bottone
M G
MG
Danova
M
M
Pellicciari
C
C
D016475:3T3 Cells; D000818:Animals; D001943:Breast Neoplasms; D002453:Cell Cycle / Q000502:physiology*; D002460:Cell Line; D004247:DNA / Q000032:analysis / Q000378:metabolism*; D004273:DNA, Neoplasm / Q000032:analysis / Q000378:metabolism*; D005260:Female; D005347:Fibroblasts; D005434:Flow Cytometry / Q000379:methods; D005455:Fluorescent Antibody Technique; D005910:Glioma; D006801:Humans; D019394:Ki-67 Antigen / Q000032:analysis; D046911:Macromolecular Substances; D008546:Melanoma, Experimental; D051379:Mice; D018809:Proliferating Cell Nuclear Antigen / Q000032:analysis; D051381:Rats; D012264:Ribonucleotide Reductases / Q000032:analysis*; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D019394:Ki-67 Antigen; D046911:Macromolecular Substances; D018809:Proliferating Cell Nuclear Antigen; D004247:DNA; D012264:Ribonucleotide Reductases