DNA flow cytometry

Hedley, D. W., Friedlander, M. L., and Taylor, I. W. (1985) Application of DNA flow cytometry to paraffin-embedded archival material for the study of aneup-loidy and its clinical significance. Cytometry 6, 327-333. 

Kangasniemi M, Veromaa T, Kulmala J, Parvinen M, Toppari J (1990) DNA flow cytometry of defined stages of rat seminiferous epithelium effects of 3 Gy of high energy x-irradiation. J Androl, 11 312-317. 

BrdU/DNA flow cytometry offers flexibility and diversity in the study of cell kinetics from cells in culture to human tumors in vivo. The essence of the procedure is to pulse label with BrdU by a short-term incubation in vitro or by a single injection in vivo samples are then taken at time intervals thereafter and stained after fixation in ethanol. The cells are then stained with a monoclonal antibody against BrdU that can be either directly conjugated to a fluoro-chrome (usually fluorescein isothiocyanate [FITC]) or, alternatively, bound to a second antibody conjugated with FITC. The cells are then counterstained with propidium iodide (PI) to measure the DNA content and analyzed on the flow cytometer. The results are displayed as linear-red fluorescence on the x-axis vs linear or log-green fluorescence on they-axis. 

BUdR/DNA flow cytometry offers flexibility and diversity in the study of cell kinedcs from cells in culture up to human tumors in vivo. Developments in monoclonals, staining techniques, and analysis of data are condnually improving the sensidvity of the technique that is now replacing the outdated HTdR/autoradiography procedure in many laboratories. 

Orndal C, Carlen B, Akerman M, et al. Chromosomal abnormality t(9 22)(q22 ql2) in an extraskeletal myxoid chondrosarcoma characterized by fine needle aspiration cytology, electron microscopy, immunohistochemistry and DNA flow cytometry. Cytopathology. 1991 2 261-270. 

Ben-Izhak B, Levy R, Weill S, et al. Anorectal malignant melanoma a clinicopathology study, including immunohistochemistry and DNA flow cytometry. Cancer. 1997 79 18-25. 

DNA flow cytometry is a very useful tool that permits rapid, objective assessment of a large number of cells but may not be readily available. Comet assay, when combined with centrifugal elutriation, can provide a useful in vitro model to study differences in metabolism and the susceptibility of different te,sticular cell types to DNA-damaging compounds. Thus, new findings using these systems should lead to greater knowledge about why a chemical or class of chemicals can cause testicular toxicity. 

Schell H et al (1991) Cell cycle kinetics of human anagen scalp hair bulbs in thyroid disorders determined by DNA flow cytometry. Dermatologica 182(l) 23-26... 

Decensi A, Bruno S, Costantini M, Torrisi R, Curotto A, Gatteschi B, Nicolo G, Polizzi A, Perloff M, Malone WF et al (1994) Phase Ila study of fenretinide in superficial bladder cancer, using DNA flow cytometry as an intermediate end point. J Nat Cancer Inst 86 138-140... 

Solanum aviculare 250-ml shake flasks 100 - 600 rpm (3.5 cm throw) t 10 d cellular DNA distribution (flow cytometry) [110]... 

A variation on this method, called fluorescent in situ hybridization (FISH), uses fluorescent-labeled DNA and RNA probes for detection and visualization of single cells by microscopy or flow cytometry.7 80 The FISH technique is popular because of its sensitivity and speed of visualization fluorescent dyes can be used to produce probes with different colors for simultaneous detection of several organisms.76,81,82... 

One of the first applications developed for flow cytometry was cell cycle analysis.2 There are numerous intercalating fluorescent DNA and RNA staining reagents that can be used to determine the amount of DNA in cells, an indicator of cell cycle stage and progression, as demonstrated in Figure 7.3. Nucleic acid dyes may be selective for DNA... 

Lowe M, Spiro A, Zhang YZ et al (2004) Multiplexed, particle-based detection of DNA using flow cytometry with 3DNA dendrimers for signal amplification. Cytometry Part A 60A 135-144... 

The ability of some fluorescent dyes to bind DNA quantitatively is exploited in flow cytometry to determine the DNA content of a cell. Dyes such as propidium iodide that bind double-stranded DNA stoichiometrically can be used for the purpose. The intensity of red fluorescence is directly related to the amount of DNA bound by propidium iodide. By comparing the fluorescence intensity of the test specimen and, in turn, its DNA content to the fluorescence intensity of specimens containing normal diploid amounts of DNA, a DNA histogram can be generated. By computing a DNA index, which is the ratio of DNA content of a test specimen to the DNA content of a specimen containing a normal diploid population, information related to the presence of an aneuploid tumor population can be obtained. The DNA index of 1 would imply that the DNA in the test specimen is from a normal diploid population (2N DNA), whereas the DNA index of an aneuploid population will be greater or less than 1. Thus, the DNA index of a tetraploid (4N DNA) would be 2. 

Likewise, the determination of DNA index by flow cytometry and the ability to assess the fraction of cells in the S phase also have applications in oncology, as was discussed earlier. 

It can be concluded from the data that RP-HPLC, together with other techniques such as microscopy, flow cytometry and DNA analysis, can contribute to the determination of the effect of salinity on the diversity of phytoplankton [284],... 

M. Estrada, P. Henriksen, J.M. Gasol, E.O. Casamayor and C. Pedros-Alio, Diversity of planktonic photoautotrophic microorganisms along a salinity gradient as depicted by microscopy, flow cytometry, pigment analysis and DNA-based methods. FEMS Microbiol. Ecol. 49 (2004) 281-293. 

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