Flow microfluorometric analysis of cellular DNA: Critical comparison of mithramycin and propidium iodide.

Hamilton, Valerie T.; Habbersett, Mary Cassidy; Herman, Chester J.

doi:10.1177/28.10.6448270

Cited by 36 publications

(16 citation statements)

References 11 publications

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“…the decrease in fluorescent intensities of PI-stained lymphocytes from pristane-treated rats did not appear to be due to a decrease in double-stranded RNA content. It should also be noted that these results were in agreement with numerous reports which indicate that the contribution of double-stranded RNA in lymphocytes stained with the DNA intercalating dyes is relatively negligible [24,[26][27][28][29].…”

Section: Resultssupporting

confidence: 92%

“…Controls were cells isolated from PBS or heptadecane injected rats. [25][26][27] had no effect on the observed staining, characteristics; i.e. the decrease in fluorescent intensities of PI-stained lymphocytes from pristane-treated rats did not appear to be due to a decrease in double-stranded RNA content.…”

Section: Resultsmentioning

confidence: 47%

“…Third, RNase treatments did not affect the PI-staining charac-teristics of lymphocytes from normal or pristanetreated rats, indicating that the observed effects of pristane were not attributable to differences in double-stranded RNA content. It should also be noted that although double-stranded RNA may contribute to the staining properties of certain cell types, this reportedly is not a factor with small lymphocytes [24,[26][27][28][29]. Fourth, the pH of the fixative solution was also critical, since significant differences in staining characteristics were observed only when the cells were fixed at a physiological pH prior to PI staining (see Fig.…”

Section: Discussionmentioning

confidence: 93%

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Changes in the DNA of lymphocytes from pristance treated rats

et al. 1987

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The effects of pristane (2,6,10,14-tetramethylpentadecane) on the cellular DNA of lymphoid cells from Copenhagen rats were examined by flow cytometry. Significant reductions in the mean relative fluorescent intensities of propidium iodide (PI) stained lymphocytes from peripheral blood, spleen, thymus and lymph nodes were observed after a single intraperitoneal injection of pristane. The altered PI staining characteristics were observed as early as 4 days and reached a maximum decrease between 1-4 weeks (depending upon the lymphoid cells examined) post pristane treatment. The pristane-induced effects on peripheral blood lymphocytes were observed to be dose dependent, transient and reinducible by a subsequent exposure to pristane. Further analyses, using gas-liquid chromatography to detect pristane in the blood and lymphoid tissues of treated rats, indicated significant increases over normal amounts of pristane. Furthermore, correlations existed between the times of maximum decrease in the fluorescence of PI stained cells and the amounts of pristane detected within the respective lymphoid tissues. By contrast no changes in the PI staining characteristics of kidney cells were observed, even though appreciable amounts of pristane were detected in this organ. Diphenylamine analyses indicated no differences in the amounts of DNA in lymphoid cells from pristane treated and untreated rats. Furthermore, lymphocytes from pristane-treated rats did not exhibit decreased fluorescence when fixed at pH 10 rather than pH 7.4 prior to PI staining. Collectively these results suggest that pristane may preferentially induce qualitative rather than quantitative changes in the DNA of lymphocytes.

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Section: Resultssupporting

confidence: 92%

Section: Resultsmentioning

confidence: 47%

Section: Discussionmentioning

confidence: 93%

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Changes in the DNA of lymphocytes from pristance treated rats

et al. 1987

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“…All aliquots were fixed in 50% ethanol (-20°C) and stored at 4°C until analysis. To obtain a fluorescent-stained single cell suspension, the staining procedure was carried out in the dark at room temperature for 30 min as previously described by Hamilton, Habbersett & Herman (1980). Briefly, 1 ml of cell suspension (about 3 106 cells) was centri¬ fuged for 10 min (360 g) and the ethanol discarded.…”

Section: Flow Cytometrymentioning

confidence: 99%

Effects of a single injection of a new depot formulation of an LH-releasing hormone agonist on spermatogenesis in adult rats

Kroonenburgh¹,

Beck²,

Vemer³

et al. 1986

Journal of Endocrinology

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Adult male Wistar rats were treated with a single injection (500 micrograms s.c.) of a new biodegradable depot formulation of the LH-releasing hormone (LHRH) analogue [D-Ser(But)6]AzGly10-LH-RH (Zoladex; ICI 118,630) to evaluate its potential for inhibiting spermatogenesis. The drug produced a marked (P less than or equal to 0.05) decrease in serum concentrations of FSH, LH and testosterone with a maximum effect 14 days after treatment. Since striking focal histological changes were seen in the testis after only 1 week, at a time when changes in serum gonadotrophins were minimal, there may be a direct effect of the LHRH analogue on spermatogenesis. Degenerative changes in germ cells as well as Sertoli cells could be observed. Flow-cytometric analysis of testicular cell suspensions showed a significant decline in the absolute numbers of haploid cells (spermatids), tetraploid cells (mainly pachytene spermatocytes) and of the numbers of cells in the S-phase of the cell cycle. This suggests that the drug also inhibits proliferation of spermatogonia and/or primary spermatocytes. Testis weight, serum hormone concentrations, and histological and cytological parameters returned to essentially normal values 52 days after the injection. It is concluded that this new method of administration may have practical and pharmacokinetic advantages for the purpose of reversible inhibition of spermatogenesis.

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“…For histological measurement, the germ cell maturation sequence of 14 stages as proposed by Leblond & Clermont (1952) (-20°C) and stored at 4°C until analysed. Staining was carried out as described elsewhere (Tannenbaum, Cassidy, Alabaster & Herman, 1978;Hamilton, Habbersett & Herman, 1980). Briefly, 1 ml cell suspension (about 3 106 cells) was centrifuged for 10 min at 370g and the ethanol was discarded.…”

Section: Methodsmentioning

confidence: 99%