Analysis of flow cytometric data of irradiated cell populations

Abstract

Flow cytometry (FCM) and autoradiography have been applied to determine changes in the cell kinetics of irradiated cells. Synchronized L-929 cells were irradiated with 10 Gy of X-rays when progressing from B1- to S-phase of the cell cycle. In this study three methods to analyse DNA histograms were tested for applicability on FCM data obtained from cell populations blocked or retarded in the cycle: A) the Gaussian integral method, B) the peak-half-reflection method, and C) the rectangle method. Since histograms from synchronized cells are heavily distorted as compared to those obtained from exponentially growing cells and are quite similar to histograms from irradiated cells, they were used to test the suitability of the evaluation methods. Comparing the evaluated FCM data with the autoradiographic results from the same experimental series, the Gaussian integral method proved to be superior to the two other relatively simple approximation methods. The FCM histograms of irradiated cells were therefore analyzed only by the Gaussian integral method. It was shown that a considerable fraction of cells is still in the S-phase 25 h post irradiation, the DNA synthesis of which had ceased, as shown by autoradiography. This indicated that parallel measurements using FCM and autoradiography yield additional information on cell kinetic changes that cannot be obtained from applying one of the two methods used.