A comparison of mathematical methods for the analysis of DNA histograms obtained by flow cytometry

Abstract

Twelve methods for analysing FCM-histograms were compared using the same set of data. Some of the histograms that were analysed were simulated by computer and some were taken from experiments. Simulated data were generated assuming asynchronously growing cell populations and (i) measurement coefficients of variation (CV) from 2 to 16%; (ii) constant measurement CV or CV's increasing from G, to G , phase, and (ii) varying fractions of cells in each phase. Simulated data were also generated assuming synchronous cell populations i n which a block in early S phase was applied and released. DNA histograms were measured for L-929 cells at various times after mitotic selection. Labelling indices were also measured for these cells at the same time.The fractions of cells in the G,, S, and (G2 + M) phases were calculated by each analytical method and compared with the actual fractions used for simulation, or in case of exp&rimental data., with autoradiographic results. Generally, all methods yielded reasonably accurate fractions of cells in each phase with relative errors in the range of 1&20%. However, most methods tended to overestimate G, fractions and underestimate S fractions. In addition, variations in the shape of the S phase distribution often caused considerable errors. Phase fractions were also calculated for histograms of kinetically perturbed populations, simulated as well as experimental The errors were only slightly larger than for histograms from asynchronously growing cell populations.Several methods for estimating cell-cycle-traverse parameters of DNA histograms generated from flow cytometry (FCM) have been published in the last few years (Dean

Proliferation kinetics of perturbed cell populations determined by the bromodeoxyuridine‐33258 technique: Radiotoxic effects of incorporated [3H]thymidine

“…The flow cytometric measurements were performed using a Phywe ICP 22 detector, However, for the present analysis only the fractions of cells in G2 + M were required this fraction was determined by a computer program that assumes a rectangular shape for the distribution of S cells (2). This method was selected because the more sophisticated evaluation procedures did not prove to be more accurate, as shown in an extended study by Baisch et al (1). on the left-hand side of the G1 peak; and the last two histograms, 10 and 16 hr, show an increasing number of the original G1 cells after mitosis.…”

DNA ploidy and proliferative activity (S-phase) in childhood soft-tissue sarcomas: their value as prognostic indicators

“…Represen used as an index of proliferative activity of the tumour. The S-phase fraction was calculated with the model of multiple broadened rectangles or dual cycling populations (Baish et al, 1982;Scott et al, 1992). Flow cytometric analysis disclosed the presence of four distinct categories of cellular DNA content (Figure 1).…”

A practical graphical method for estimating the fraction of cells in S in DNA histograms from clinical tumor samples containing aneuploid cell populations

“…However, one can evaluate the results obtained by the trapezoidal method by comparing them (12), for histograms with CVs in the range of 0 to 5% (1). Since all of our DNA histograms had CVs of 4% or less, we have used the Jett method as a comparative reference for the trapezoidal graphical method in the analysis of real DNA histograms.…”