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At least 55 records · Page 3Linked to original sources
PubMed1979

Multi-user system for analysis of data from flow cytometry.

A new program is described for the analysis of DNA histograms from flow cytometry. The fundamental model representing the cell population is similar to one described previously. It assumes the population is grouped into compartments, each consisting of cells having approximately the same DNA content. After staining the cells with an appropriate fluorochrome, the fluorescence distribution of cells within each compartment is assumed to be Gaussian. In the present algorithm, the parameters of the model can either be computed directly by the program from the data, or can be specified as input by the user. When synchronous cell populations lacking distinct G1 and G2/M phases are analyzed, the parameter values must first be obtained using an appropriate control. Percentages of cells in the various compartments are computed using a gradient search method described by Bevington.

Computers↗
PubMed1992

[Study of the pisum sativum agglutinin receptors and DNA ploidy during carcinogenesis in esophageal epithelium by flow cytometry].

The relative mean contents of pisum sativum agglutinin receptors and DNA of human normal esophageal epithelium, its atypical hyperplasia as well as esophageal carcinoma were determined respectively by flow cytometry. The results revealed that the relative mean contents of pisum sativum agglutinin receptors which denote membrane composition and DNA which denote nuclear composition were increased gradually from normal, dysplastic and then malignant esophageal tissues. This suggested that during the neoplastic transformation, changes of the cell membrane component may correlate with changes of the cell nuclear component.

Aneuploidy↗
PubMed1979

Fluorescent DNA probes for flow cytometry. Considerations and prospects.

Techniques employing base specific deoxyribonucleic acid (DNA)-binding fluorochromes and flow cytometry (FCM) are potentially useful for obtaining information of the compositional features of chromatin or chromosomes of mammalian cells. Fluorescent compounds which form complexes preferentially at the A-T rich regions (i.e., DNA-reactive Hoechst dyes) or the G-C rich regions (i.e., mithramycin, chromomycin, olivomycin) in DNA are available and compatible with current FCM technology as are other compounds (i.e., ethidium bromide, propidium iodide) which show little or no base specificity and bind by intercalation in the double stranded regions of helical DNA. Energy transfer between appropriate DNA-bound dyes is a reflection of the quantity and proximity of regions containing the respective base pair segments. Since extrinsic fluorescent probes provide only a measure of available binding sites or regions unobstructed by chromatin-associated or chromosomal-associated proteins, interpretations of fluorescence measurements need to be substantiated by adequate control measures.

Animals↗
PubMed1992

Studies with biotinylated RBC: (1) use of flow cytometry to determine posttransfusion survival and (2) isolation using streptavidin conjugated magnetic beads.

Methods are reported for the quantitation and isolation of biotinylated red blood cells (B- RBC). The first method is for determination of posttransfusional survival of rabbit RBC by flow cytometry. The survival of B-RBC was measured using both fresh and paraformaldehyde-fixed cells with similar results. The posttransfusion survival of rabbit RBC measured in this way was normal. There was no indication of increased cell destruction due to antibodies directed against B-RBC and no evidence for loss of biotin from circulating cells. The second methodology is for the isolation of B-RBC from blood with streptavidin-coated magnetic beads. At least eighty percent of positive cells were recovered with very few false positives. Both methods may be helpful in the study of resealed erythrocytes.

Animals↗
PubMed1992

Cell cycle analysis of asynchronous cell populations by flow cytometry using bromodeoxyuridine label and Hoechst-propidium iodide stain.

Continuous labelling of cells with deoxybromouridine (BrdUrd) followed by staining with a bis-benzimidazole (Hoechst 33258) and a phenanthridinium (propidium iodide or ethidium bromide) allows the cells to be separated by flow cytometry according to the extent of their DNA replication. This BrdUrd-Hoechst/PI method has been used mainly to observe perturbations of the cell cycle in synchronously growing cells. In this paper we demonstrate that, when the method is applied to asynchronously dividing cells, more extensive information can be derived about the effects of cytotoxic and other treatments on the kinetics of the cell cycle. The interpretation of the data is explained, the effects of different types of cytotoxic agent are described, and the method is compared briefly to other methods for following cell cycle kinetics.

Animals↗
PubMed1979

Cell-cycle distribution of urothelial tumour cells as measured by flow cytometry.

The fraction of cells in S + G2 + mitosis from 54 urothelial tumours was calculated by flow cytometry after acridine orange (AO) staining of cells obtained by bladder irrigation or biopsy. Fluorescence signals emitted by the AO-stained DNA and RNA of each cell were separated optically and measured for 5,000 cells per specimen. The patients were classified by the histology of their tumours and clinical data into 5 diagnostic categories: NED (no evidence of disease, but history of bladder tumour), 3; papilloma, 8; non-invasive papillary carcinoma, 8; carcinoma in situ, 17 and invasive carcinoma, 18. The fraction of cells with DNA values in S + G2 + M of the cell cycle varied between 7 and 57% of the total, with a wide range within each diagnostic category, but no statistically significant differences between the groups. The proportion of cells in S + G2 + M from an individual tumour was not correlated with histologic grade or clinical behaviour. The possibility that some tumour cells with DNA values above G1 level are quiescent cells arrested at S or G2 is discussed.

Cell Count↗
PubMed1979

Flow cytometry of human gynecologic specimens using log chromomycin A3 fluorescence and log 90 degrees light scatter.

Flow cytometry and electronic cell sorting are being investigated to screen gynecologic specimens for cervical neoplasia. Cellular DNA content is quantitated by Chromomycin A3 fluorescence and cell size is quantitated by 90 degrees light scatter; the logarithms of the measured intensities are used to produce a two parameter histogram. To determine the cell types responsible for signals in various histogram regions, systematic electronic cell sorting is performed. The sorted fractions are sedimented into microscope slides and stained by the Papanicolaou technique. The cells in each fraction are identified by conventional cytomorphologic criteria. Morphologic analysis of sorted cells reveals histogram regions corresponding to specific cell types. One very important region contains the highest concentration of signals from abnormal cells and is therefore the best region to analyze for specimen abnormality. However, because a significant number of signals in this region are from normal cells, specimens cannot be diagnosed by their analysis. Another important histogram region is composed primarily of signals from endocervical columnar and metaplastic cells. The presence of such cells is a good criterion for specimen adequacy, therefore analysis of signals in this region is essential to assess specimen adequacy for automatic screening.

Cervix Uteri↗
PubMed1979

Lymph node reactivity to experimental bladder tumor in preimmunized animals as measured by two parameter flow cytometry.

Lymph node lymphocyte reaction to an explanted, transplantable mouse bladder tumor (MBT-2) was investigated by flow cytometry in animals previously immunized with irradiated tumor cells. Nodal lymphocytes in representative samples from four different lymph node sites were differentially stained for DNA and RNA with the fluorescent dye acridine orange; cell proliferation and the increase in RNA content were measured. Immunization abrogated tumor growth; one immunization reduced tumor take to 25 per cent of the animals, and two immunizations to 14 per cent. Lymphocyte reactivity to the tumor was reflected both by an increase of DNA synthesizing cells and by diploid cells with high RNA. The latter response was more pronounced and thus the more sensitive parameter for measuring immunologic lymph node reactivity. The juxtatumoral node displayed the most pronounced reactivity, but all node sites showed some degree of reaction.

Animals↗
PubMedDate not supplied

Flow cytometry to evaluate the parasitemia of Plasmodium falciparum.

The resistance of plasmodiums to the current antimalarial agents has spurred the search for new active molecules of vegetal origin or chemical synthesis. The screening of antimalarial molecules "in vitro" was done by simple but tedious techniques such as quantification of parasitemia through optical microscopy or by using radioactive markers. We have developed a new method to evaluate parasitemia by using the ODAM ATC 3000 flow cytometer and biological cell sorter. We selected the ethidium bromide for labelling the nucleic acids of Plasmodium falciparum, and we optimised the method by using a mathematical model: the design of Hadamar. This simple technique presents the advantage of being an objective and rapid count of large number of red cells (10(6) x 10(7)). This method is rapid, reliable, reproducible, devoid of subjectivity and provides more precise results than those of optical microscopy. The good correlation between paraitemia measured by optical microscopy and fluorescence obtained by flow cytometry allows us to recommend this technic for the screening of new antimalarial molecules.

Ethidium↗
PubMed1979

Measurement and purification of human chromosomes by flow cytometry and sorting.

The 24 human chromosome types of normal diploid fibroblast cell strain were classified into 15 groups by high-resolution flow cytometry on the basis of 33258 Hoechst fluorescence. Chromosomes associated with each group were flow sorted onto microscope slides and identified by quinacrine banding analysis. DNA cytophotometry of metaphase chromosomes from the same cell strain supported and extended this identification. Four of the groups purified were due to chromosomes of a single type--namely, chromosomes 5, 6, 13, and 17. Eight additional groups were also separated and found to contain the following chromosomes: 1 and 2; 3 and 4; 7, 8, and X; 9--12; 14 and 15; 16 and 18; 20 and Y; and 19, 21, and 22. The average purity for the 12 sorted fractions was 78%.

Cell Line↗
PubMed1978

Characterization of bladder tumours by flow cytometry on bladder washings.

The DNA content of individual nuclei in bladder washings was estimated by means of flow cytometry. A method of preparation using a detergent to produce single nuclei was used. By means of a differential count the DNA distribution was corrected for the content of leucocytes. The DNA histograms were divided into a 2c fraction containing nuclei with less than 3c DNA, a 4c fraction with 3c--6c DNA and an 8c fraction with more than 6c DNA. Three groups of patients were investigated: 17 patients with invasive bladder tumours, 38 patients with non-invasive bladder tumours, and 22 patients with normal bladders. The washings from patients with invasive tumours contained significantly larger 4c and 8c fractions than washings from normal bladders. There was a correlation between cell anaplasia and the DNA distribution: grade III bladder tumours had larger 4c and 8c fractions than grade II tumours.

Cell Nucleus↗
PubMed1979

Size measurements on isolated rat heart cells using Coulter analysis and light scatter flow cytometry.

Isolated ventricular muscle cells from the adult rat heart have been examined by both Coulter analysis and light scatter flow cytometry. The dispersed cell preparations contain two main cell types: viable, rod-shaped cells and damaged, round cells. Coulter analytical techniques provided statistical data on cell volume for both cell types. The contribution of each population to the Coulter pulse height distributions were separated by a subtraction method using data obtained from digitonin-treated preparations that contain only round cells. A shape factor for cells aligned with the flow direction was computed from light microscope measurements and the effects of cell orientation within the Coulter aperture were approximately assessed. The estimated volumes for intact myocytes compare favourably with those reported in the literature. No significant size difference was observed between fresh and fixed cells. Narrow angle, forward light scatter measurements were made on individual cells flowing across a focused laser beam. Both scatter pulse height and pulse width (pulse duration) distributions were collected. Values for myocyte length calculated from pulse width information agree well with published data and confirm that the hydrodynamic forces in the flow system produced alignment of the cells with the flow direction. Scatter pulse width distributions reveal two distinct peaks assignable to either rod or round cells. Preliminary electronic gating experiments, using pulse height signals, suggest that signals derived from round cells could be eliminated entirely using a gating regime based on pulse width. This would enable flow cytometric measurements to be made on only the intact myocytes present in heterogeneous preparations.

Animals↗
PubMed1992

Alteration of intracellular DNA and RNA patterns by liver arginase studied with flow cytometry.

Previous study of lymphocyte proliferation in the presence of liver arginase has indicated that arginine-depletion in the culture medium plays an important role in inhibiting cell proliferation. The inhibitory effect of both liver arginase and arginine-free condition on DNA, RNA and protein syntheses in PHA-stimulated lymphocytes were studied in cultures by measuring the incorporation of labeled precursors. Simultaneously, their influence on DNA and RNA contents in cells stained by acridine orange was investigated by automated flow cytometry. With 3 micrograms/ml arginase, the syntheses of DNA, RNA and protein were markedly inhibited after 72 h of culture. The degrees of inhibition were close to that induced in an arginine-free condition. The DNA and RNA contents of the individual cell, either cultured with 3 micrograms/ml arginase or in arginine-free medium, were arrested in G0/G1 phase. The results of cell arrest in G0/G1 phase were similar whether the cells were cultured for 24, 48 or 72 h.

Animals↗
PubMed1992

[Prognostic significance of nuclear DNA measurement with flow cytometry for laryngeal squamous cell carcinomas].

DNA ploidy, S% and proliferative index (PI) of carcinomatous cell were evaluated with FCM in 41 patients with laryngeal primary squamous cell carcinomas which were operated on in our department during the period between Jan. 1979 to Oct. 1987. The results showed that DNA aneuploidy was identified in 65.85% of the cases (27/41) and patients with aneuploid carcinomas had higher S% (mean +/- standard: 23.22 +/- 9.57%), higher PI (mean +/- standard: 37.59 +/- 9.57%) and higher rate of relapse and/or metastasis than those with diploid tumours (55.55% vs 11.77%, P < 0.025). Patients with aneuploid carcinomas also correlated with an unfavorable prognosis compared with those with diploid tumors. Cases with aneuploid and diploid carcinomas had 66.11% and 92.86% respectively for two years postoperation survival rate; 46.74% and 92.86% respectively for five years postoperation survival rate (Log-rank test: chi 2 = 5.96, P < 0.025). We conclude that nuclear DNA measurement with flow cytometry is of important significance in estimating prognosis and guiding treatment for laryngeal squamous cell carcinomas.

Aged↗
PubMed1979

Regional lymph node reactivity in explanted bladder cancer of mice as measured by flow cytometry.

The reactivity of lymphocytes in lymph nodes draining the site of a transplantable experimental bladder tumor (MBT2 in C3H/HeJ mice) has been measured in a multiparameter flow cytometry system. Acridine orange was used as a nucleic acid probe. This dye intercalates in helical DNA, emitting green (530 nm) fluorescence upon exposure to blue light; it stacks to single-stranded RNA, emitting red (640 nm) fluorescence. The relative magnitude of the increase of lymphocyte DNA and RNA has been evaluated simultaneously in tumor-draining nodes, in nondraining nodes of the same animal, and in untreated control animals. Stimulation of the regional node lymphocytes could be observed after 20 days but not after 10 days. It was uniformly high at 35 days. The transcriptive response (increased proportion of lymphocytes with high RNA) was more pronounced than the proliferative (increased proportion of lymphocytes with more than diploid DNA). The histological changes in the stimulated nodes resembled closely those described by others in human tumor-draining nodes. The described method has the advantage of being simple, rapid, and able to measure a representative part of the whole-cell population.

Animals↗
PubMed1979

Flow cytometry analysis of early DNA content changes in human and monkey cells following infection with Simian Virus 40.

Simian virus 40 (SV40) is capable of inducing cellular DNA synthesis in permissive and nonpermissive cells. Utilizing flow cytometry, we analyzed the DNA content changes in two diploid human cell strains and two monkey cell lines. The osteogenesis imperfecta (OI) human skin fibroblasts were induced into DNA synthesis, and within one to two cell generations, a polyploid cell population was produced. With WI-38 phase II cells, a similar pattern of increased cycling of cells into DNA synthesis was observed; however, the majority (approximately 60%) of the cells were blocked in the G2 + M phase of the cell cycle. At later time intervals, an increase in the G1 population was demonstrated. The two monkey cell lines responded to SV40 virus with an accumulation of cells in the G2 + M phase of the cell cycle. Thus, two diploid human cell strains exhibited different cell cycle kinetics early after infection with SV40 virus. The one strain (WI-38) behaved similarly to the two monkey cell lines studied. The other strain (OI) responded similarly to nonpermissive (transforming) cells infected with SV40 virus.

Animals↗
PubMed1979

Slit-scan flow cytometry of mammalian chromosomes.

A flow cytometer has been constructed which measures total fluorescence and the distribution of fluorescence along isolated, stained mammalian chromosomes. In this device, chromosomes flow lengthwise at 4 m/sec through a 1-micrometer thick laser beam. The fluorescence from each chromosome is recorded at 10 nsec intervals; the sequence of recorded values represents the distribution of fluorescence along the chromosome and is stored in the memory of a waveform recorder. The total fluorescence of each chromosome is also measured and recorded. Preliminary studies show that doublets of 1.83 micrometers diameter microspheres flow with their long axes parallel to the direction of flow and that the two microspheres are resolved in the slit-scan profile. Ethidium bromide stained Muntjac and Chinese hamster chromosomes have also been slit-scanned. Centromeres were resolved in many of the Nos. 1 and 2 Chinese hamster chromosomes and the Nos. 1 and X + 3 Muntjac chromosomes.

Animals↗