Rate and Time of Dna Synthesis of Individual Chinese Hamster Cells

Dormer, Peter G.; Brinkmann, W; Born, R; Steel, G. Gordon

doi:10.1111/j.1365-2184.1975.tb01228.x

Cited by 26 publications

(13 citation statements)

References 17 publications

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“…The two obtained densities agree well and show that there are more cells in the beginning and end of the S phase, implying that the total rate of DNA replication is slower and cells spend more time in these parts than in the middle of the phase. This result coincides with what has been shown before (Larsson Bertuzzi et al, 1984;Dörmer et al, 1975). Assuming a specific value for the parameter b governing the variance of T S , (6) provides an estimate of the reference replication rate v(x) from the estimated DNA density.…”

Section: Results For the Mcf-7 Cell Linesupporting

confidence: 84%

“…The only distribution of interest in this paper is that of the S phase duration. This distribution is not known, although it is believed to be skewed with a tail toward higher values of T S (Baisch and Otto, 1993;Dörmer et al, 1975). Baisch and Otto (1993) considered a log normal distribution, whereas Macdonald (1981) suggested a gamma distribution; for computational reasons we adopt the latter.…”

Section: Model Assumptionsmentioning

confidence: 97%

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Estimating the Total Rate of DNA Replication Using Branching Processes

et al. 2008

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Increasing the knowledge of various cell cycle kinetic parameters, such as the length of the cell cycle and its different phases, is of considerable importance for several purposes including tumor diagnostics and treatment in clinical health care and a deepened understanding of tumor growth mechanisms. Of particular interest as a prognostic factor in different cancer forms is the S phase, during which DNA is replicated. In the present paper, we estimate the DNA replication rate and the S phase length from bromodeoxyuridine-DNA flow cytometry data. The mathematical analysis is based on a branching process model, paired with an assumed gamma distribution for the S phase duration, with which the DNA distribution of S phase cells can be expressed in terms of the DNA replication rate. Flow cytometry data typically contains rather large measurement variations, however, and we employ nonparametric deconvolution to estimate the underlying DNA distribution of S phase cells; an estimate of the DNA replication rate is then provided by this distribution and the mathematical model.

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Section: Results For the Mcf-7 Cell Linesupporting

confidence: 84%

Section: Model Assumptionsmentioning

confidence: 97%

Estimating the Total Rate of DNA Replication Using Branching Processes

et al. 2008

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“…In Table 1, only the results of Straus are from in vivo administration of tritiated thymidine. In vitro measurement has obvious advantages, and permits use of FUdR to enhance labeling (11,12). Thin slices cut at less than 1 mm freehand are better than minced tissue because of less trauma and absence of fragmentation.…”

Section: Thymidine Labeling Indexmentioning

confidence: 99%

Practical breast carcinoma cell kinetics: Review and update

Meyer

McDivitt

Stone

et al. 1984

Breast Cancer Res Tr

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The S-phase fraction (SP) measured by flow cytometry of DNA and the thymidine labeling index (TLI) measured autoradiographically indicate the proportion of carcinoma cells currently synthesizing DNA and reflect the rate of proliferation. The TLI and SPF are lognormally distributed. The median TLI performed to maximize precursor uptake is near 5% (5 labeled carcinoma cells per 100) the mean near 7%, and the range from less than 1% to near 40%. Corresponding values for the SPF measured by DNA flow cytometry are slightly higher when appropriate measures are taken to reduce background debris counts and other artefacts. Residual elevation of SPF above TLI may result from S-phase arrested cells. Flow cytometric histograms show that clearly aneuploid cell lines exist in 50-80% of primary breast carcinomas. Aneuploid breast carcinomas have higher mean TLI than diploid breast carcinomas, and therefore proliferate more rapidly. They also more frequently lack estrogen receptor (ER). Carcinomas with minimal nuclear anaplasia, particularly those of tubular, mucinous, infiltrating lobular and adenocystic types have low TLI and SPF, whereas carcinomas with highly anaplastic nuclei, including medullary carcinomas, have high TLI and SPF. TLI and SPF correlate inversely with ER and PgR content, have no relationship to axillary lymph nodal status, and have a weak positive correlation with tumor size and a weak negative correlation with age. High TLI predicts a high risk of early relapse after primary therapy for both node-negative and node-positive carcinomas. Carcinomas that produce brain metastases have particularly high TLI. Current evidence suggests that high SPF and aneuploidy may prove to have prognostic significance like TLI.

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“…In addition, an intracellular precursor pool of dThd monophosphate exists at the beginning of BrdUrd incorporation (7) and should not mix with the newly synthesized BrdUrd monophosphate. Dormer applied fluorodeoxyuridine (FdUrd) to block the endogenous synthesis of dThd monophosphate for use in quantitative 14C-autoradiography (5,11,13). He first preincubated the DNA synthesizing cells for 6 min with FdUrd to clear the endogenous dThd monophosphate pool before incubating the cells with 14C-dThd.…”

mentioning

confidence: 99%

Effect of 5‐fluoro‐2′‐deoxyuridine (FdUrd) on 5‐bromo‐2′‐deoxyuridine (BrdUrd) incorporation into DNA, measured with a monoclonal BrdUrd antibody and by the BrdUrd/hoechst quenching effect

Ellwart

Dormer

1985

Cytometry

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The purpose of this study was to improve the application of bromodeoxyuridine (BrdUrd) for the flow cytometric analysis of cell kinetics. In order to obtain a quantitative measure of the DNA synthesis rate (or the number of divided cells), BrdUrd should replace thymidine (dThd) completely in the newly synthesized DNA strands. The de novo synthesis of dThd monophosphate competing with BrdUrd incorporation was stopped by fluorodeoxyuridine (FdUrd). Cells of a human leukemic cell line (REH) were exposed 4.0 BrdUrd for either 20 min, 8 h, or 24 h. Bromodeoxyuridine incorporation was determined by a monoclonal antibody as well as by the BrdUrd/Hoechst (HI technique. Counterstaining of the DNA was performed with propidium iodide or ethidium bromide. DNA fluorescences were measured in both techniques with a two-parameter flow cytometer, the histograms being analyzed by computer. It was found that FdUrd is required in the BrdUrd/ H technique for replacement of dThd at low BrdUrd concentrations and long incubation times. With short incubation periods, as used for detection by the monoclonal anti-BrdUrd antibody, FdUrd increases the incorporated BrdUrd amount when BrdUrd concentrations of 10 pM or less are applied.Key terms: Bromodeoxyuridine, fluorodeoxyuridine, DNA synthesis rate, human leukemic cell line, flow cytometry Bromodeoxyuridine (BrdUrd) is used with increasing frequency in cell kinetic studies after the introduction of the Hoechst (H) quenching effect into flow cytometry by Latt (23,241. Bohmer (2) and Kubbies and Rabinovitch (20) described methods of cell cycle analysis using the quenching of H in divided cells. Perturbations of the cell cycle of normal and tumor cells were easy to investigate with the BrdUrdM technique (1,29). Counterstaining of total DNA per cell with ethidium bromide (EB) simplified the analysis of the DNA synthesis in the first cycle (3,4,16). This two-parameter technique allows flow cytometric measurement of the DNA synthesis rate even in subcompartments of the S-phase after a few hours of BrdUrd incubation (28,151). Since it is now possible to measure DNA synthesis with the monoclonal anti-BrdUrd antibody (10,18,26), BrdUrd is being applied increasingly for cell cycle studies. It is therefore of interest to know the exact conditions of quantitative BrdUrd incorporation.The quantity of BrdUrd instead of thymidine (dThd) incorporated per unit of time into DNA may provide information on the DNA synthesis rate. For quantitative assessment of the DNA synthesis rate, it is most convenient if BrdUrd replaces dThd completely in the newly synthesized DNA strands. It is, however, to be expected that the endogenous synthesis of the DNA precursor dThd monophosphate competes with the exogenously supplied BrdUrd. In addition, an intracellular precursor pool of dThd monophosphate exists at the beginning of BrdUrd incorporation (7) and should not mix with the newly synthesized BrdUrd monophosphate. Dormer applied fluorodeoxyuridine (FdUrd) to block the endogenous synthesis of dThd monophosphate for us...

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Rate and Time of Dna Synthesis of Individual Chinese Hamster Cells

Cited by 26 publications

References 17 publications

Estimating the Total Rate of DNA Replication Using Branching Processes

Estimating the Total Rate of DNA Replication Using Branching Processes

Practical breast carcinoma cell kinetics: Review and update

Effect of 5‐fluoro‐2′‐deoxyuridine (FdUrd) on 5‐bromo‐2′‐deoxyuridine (BrdUrd) incorporation into DNA, measured with a monoclonal BrdUrd antibody and by the BrdUrd/hoechst quenching effect

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