DNA flow cytometric analysis

Figure 5.3 DNA flow cytometric analysis of cells treated with apigenin for 24 or 48 h. Each point represents the mean SD of three independent experiments. Cell cycle was monitored by a DNA flow cytometric analysis indicating as follows 0,% of G1-phase cells A,% of S-phase cells % of G2/M-phase cells. Means SD from three independent experiments are shown. p < 0.05 and p < 0.01 vs. the vehicle controls. (From Wang et at, Molecular Carcinogenesis, 28, 102-110, 2000. With permission.)...

Nuclear stains (i.e., Hoechst, DAPI) Transverse or pulse field electrophoresis Assessment of Soluble DNA Flow cytometric analysis of DNA content... 

Luna MA, el Naggar A, Parichatikanond P, et al. Basaloid squamous carcinoma of the upper aerodigestive tract. Clinicopathologic and DNA flow cytometric analysis. Cancer. 1990 66 537-542. 

Imray FP, Kidson C (1983) Perturbations of cell-cycle progression in gamma-irradiated ataxia telangiectasia and Huntington s disease cells detected by DNA flow cytometric analysis. Mutat Res 112 369-382... 

Figure 6.2 Cell cycle distributions of MCF-7 cells after treatment with TSA for 24 hours were determined by performing flow cytometric analysis. Following treatment of cells with 50 nM and 1 P-M of TSA, the samples were analyzed on a Becton-Dickinson FACStarP "" instrument and the percentage of nuclei with C], S, and C2/M DNA content was determined. p21 protein was also correspondingly increased at various intervals.

Toppari J, Bishop PC, Parker JW, Ahmad N, Girgis W, diZerega GS (1990) Cytotoxic effects of cyclophosphamide in the mouse seminiferous epithelium DNA flow cytometric and morphometric analysis. Fundam Appl Toxicol, 15 44-52. 

Larsen, J. K., Christensen, I J, Christiansen, J., and Mortensen, B. J (1991) Washless double-staining of unfixed nuclei for flow cytometric analysis of DNA and a nuclear antigen (Ki-67 or bromodeoxyuridine) Cytometry 12, 429—437... 

It is possible to permeabilize the outer membrane of normal cells (with detergent or alcohol) in order to allow propidium iodide to enter the nuclei. If we then treat the normal cells with RNase in order to ensure that any fluorescence results from their DNA content (without a contribution from double-stranded RNA), we find that the nuclei fluoresce red with an intensity that is more or less proportional to their DNA content. By the use of a red filter and a linear amplifier on the photomultiplier tube, we can detect that red fluorescence. The channel number of the fluorescence intensity will be proportional to the DNA content of the cells. The method is simple and takes about 10 minutes. Flow cytometric analysis of the red fluorescence from the particles in this preparation of nuclei from normal, nondividing cells will result in a histogram with a single, narrow peak (see the first histogram in Fig. 8.1) all the particles emit very nearly the same amount of red fluorescence. This supports our knowledge that all... 

Flow cytometric analysis of genome size requires isolating cells or nuclei in high concentrations. The nuclei are then stained with various DNA-specific fluorochromes. The stained nuclei are subsequently analyzed by a flow cytometer cell sorter. Critical to the technique is the freeing of intact cells or nuclei. 

Pollice, A A, McCoy, J. P, Jr., Shackney, S E, Smith, C A., Agarwal, J., Burholt, D R., Janocko, L. E, Hornicek, F J, Singh, S. G, and Hartsock, R. J. (1992) Sequential paraformaldehyde and methanol fixation for simultaneous flow cytometric analysis of DNA, cell-surface proteins, and intracellular proteins. Cytometry 13,432-444... 

For flow cytometric analysis of DNA content, cells under investigation, in the exponential phase of cell growth, can be treated at different concentrations... 

Yoshimtrra T, Manabe T, Imamura T, et al. Flow cytometric analysis of nuclear DNA content of duct cell carcinoma of the pancreas. Cancer. 1992 70 1069-1074. 

Blom R, Guerrieri C, Stal O, et al. Leiomyosarcoma of the uterus A clinicopathologic, DNA flow cytometric, p53, and mdm-2 analysis of 49 cases. Gynecol Oncol. 1998 68 54-61. 

Caspase 3 upregulation, cyclin A levels, P53 level. Fas ligand expression, cell size and granularity (flow cytometric analysis), annexin V binding, DNA fragmentation, nuclei condensation... 

Furthermore, the effects of several active compounds on the cell cycle of Raji cells treated with TPA were also examined by flow cytometric analysis. Generally, the cells (including cancer cells) repeats the division and the prolifiration via Gi phase (resting phase after cell division) -> S phase (DNA synthetic phase) - G2 phase (resting phase before cell... 

The frequency function for an overall population specific growth rate of 0.088 hr exhibits three modes, the first corresponding to a DNA content of 1C, the second to the most prevalent 2C population, and the final one to a population of cells containing a DNA content of 4C. The last mode is not believed to be the result of cell clumping, as all samples in these analyses were treated by sonlcatlon immediately prior to flow cytometric analysis. (The suitability of this procedure for providing accurate cell counts. 

FIGURE 7.2 Flow cytometric analysis of cell cycle distribution and apoptosis in human oral keratinocytes in response to increasing concentrations of smokeless tobacco extract. Primary keratinocytes were grown to approximately 70% confluence and then treated with various concentrations of STE for 25h. Following incubation, the cells were removed from the culture surfaces by trypsinization, and DNA content and apoptosis were determined using the method of Telford et al. Raw data were modeled using ModFit cell cycle... 

Flow Cytometric Analysis of Cell Cycle. PC-3 cells (1-2 x 10 ) were stained with propidium iodide (PI) according to a reported procedure (78). DNA content analysis was performed on Pl-stained cells using the FACSCalibur instmment (Becton Dickinson Immunocytometry System, San Jose, CA). 

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