Quantitative determination of transformed cells in a mixed population by stimultaneous fluorescence analysis of cell surface and DNA an individual cells.

Abstract

ABSTACT Cell-surface labeling with fluorescamine indicates that the fluorescence of Balb 3T3 A31 cells is considerably decreased after both viral and chemical transformation. This phenomenon, coupled with the technique of flow microfluorometry, enabled nontransformed and transformed cells to be distinguished. A Second fluorescent probe, propidium iodide, which intercalatesinto DNA, was used in combination with fluorescamine in order to obtain a ratio of cell-surface labeling to DNA content: This manipulation allowed enhanced resolution of the two op tions'and the detection of small numbers of transfor abts n tr~dominantly normal population.

Effects of 3-(3,4-Dichlorophenyl)-1,1-Dimethylurea on the Cell Cycle in Euglena gracilis

“…The propidium iodide was excited with 488 nm light, and fluorescence intensity per cell at greater than 620 nm was measured. Cells were analyzed in a FMF-77-100 flow cell (Research Developments) in a flow cytometer constructed as previously described (7,9 …”

“…The propidium iodide was excited with 488 nm light, and fluorescence intensity per cell at greater than 620 nm was measured. Cells were analyzed in a FMF-77-100 flow cell (Research Developments) in a flow cytometer constructed as previously described (7,9 Figure 1 indicates that the control cells not treated with DCMU completed S phase, G2/M, and returned to G,. Addition of DCMU at 6 h after inoculation caused cells to accumulate in the premitosis (G2/M) phase (Fig.…”

Analysis of Expression of Human Mammary Epithelial Antigens in Normal and Malignant Breast Cells at the Single Cell Level by Flow Cytofluorimetry

“…However, two important technical difficulties have hampered the use of flow cytofluorimetry in the quantitation of surface antigens on cells from solid tissue: (i) the difficulty of obtaining single cell suspensions without altering the cell surface, and thus being unable to obtain cell per cell readings of external antigens that are re moved by enzymes used in cell dissociation procedures; and (ii) the presence of different proportions of cell types other than the one to be analyzed, among the dissociated cells of the tissue, which are not distinctly different enough to be separated and obscure the specific quanti tation. In order to overcome these problems in the quantitation of anti-HME binding to breast epithelial cells, we have expanded on the proce dure developed by Hawkes and Bartholomew (8). In their procedure, they could distinguish virally or chemically transformed cells in mixed populations by doubly labeling the cells with propidium iodide, which bound to DNA, and fluorescamine, which bound differentially to normal and transformed cells (8).…”

“…In order to overcome these problems in the quantitation of anti-HME binding to breast epithelial cells, we have expanded on the proce dure developed by Hawkes and Bartholomew (8). In their procedure, they could distinguish virally or chemically transformed cells in mixed populations by doubly labeling the cells with propidium iodide, which bound to DNA, and fluorescamine, which bound differentially to normal and transformed cells (8).…”

Cell killing and mutation induction on Chinese hamster cells by photoradiations

“…In Dr. Bartholomew's laboratory, this instrument is connected to a computer system so that data of many samples could be printed out in detail. The procedures for preparing cells for FMF, the fixation and staining methods as described by Hawkes and Bartholomew (1977) Immediately after the exposure, cells were trypsinized for survival test and the remaining were transferred to a container with prewarmed medium. In split-dose experiments, prewarmed medium was added between the two conditioning doses and kept in a 37°C incubator except where different treatment was indicated.…”