Evaluation of a modeling system for S-phase estimation in XXXX by flow cytometry.

@entity1 @entity0 @entity5 @entity134 @entity44260 @entity35
Using software programs provided by Coulter Electronics, we have developed an analysis system that would address problems encountered in DNA flow cytometric analysis of heterogeneous @entity5 populations, especially where the G2-M phase of the diploid population contaminates the S-phase of the aneuploid population, causing an overestimation of cells in S phase. We used the PARA 1 and PARA 2 programs in concert and developed three analysis models: (a) for @entity5 ; (b) for @entity5 with overlapping populations; and (c) for near-diploid @entity5 . Our purpose in this paper is to determine the limits and reproducibility of this analysis system with an emphasis on @entity5 with overlapping populations. Aliquots of frozen, pulverized @entity0 tissue (50 to 100 mg), routinely used in the @entity134 receptor assay, were used for routine flow cytometric measurement of the DNA index and S-phase fraction. To determine the accuracy of the analysis when overlapping populations were present, we mixed an @entity0 cell line with @entity1 blood lymphocytes in varying ratios. @entity35 % mixture of aneuploid cells, the lowest mixture tested, still allowed analysis results within 95% confidence limits. Reproducibility of the system was assessed on frozen @entity0 tissue by intra- and interassay variation studies measuring cell cycle parameters and @entity44260 peak width. Within any sample the amount of variation (+/- 2 SD) for the G0-G1 value was +/- 4.40 for intraassay and +/- 4.60 for interassay, and the amount of variation for S phase was +/- 3.0 and +/- 3.2 for intraassay and interassay, respectively. There was no difference in the variation of estimates for G2-M (+/- 2.6 for both intra- and interassay). In this study, the @entity44260 peak greater than 5% was defined as unacceptable for accurate analysis, with the conclusion that S-phase fractions in @entity5 can be routinely analyzed in @entity1 @entity0 biopsies despite @entity5 cell heterogeneity.
