Comparative DNA analysis of solid tumors by flow cytometric and image analyses of touch imprints and flow cell suspensions

Comparative DNA analysis by flow cytometric (FCM) and image analyses (IA) has shown a high concordance rate. When present, discordance has been attributed to the presence of aneuploid cell populations detected only by IA, yet missed by FCM. This phenomenon has been explained by loss of aneuploid cells during FCM cell processing, differences in sampling area, or misinterpretation of the DNA histograms. To determine which factors are responsible for the discordance between IA and FCM, 82 fresh solid tumors from various sites were examined. Flow cytometric analysis was performed on cell suspensions isolated from the tumors, whereas IA was performed on touch imprints (IAT) and on cytosmears of the same cell suspension used for FCM (IAF). Comparison between IAT and IAF (IAT/IAF) assessed cell processing and sampling area differences, whereas IAF/FCM comparison assessed differences in apparatus and methodology as possible contributing factors to discordance. Furthermore, DNA histograms of IAT, IAF, and FCM were analyzed in the discordant cases to determine whether the discordance was due primarily to different cell populations detected (true discordance) or due to differences in histogram interpretation of the same cell populations (false discordance). IAT/IAF and IAF/FCM concordance rates (90% and 88%) were not significantly different from that of IAT/FCM (87%). False discordance accounted for most of the discordant cases in IAT/FCM comparison (six cases, 67%), whereas true discordance was seen in three cases. In all three truly discordant cases, the DNA-aneuploid cell populations detected only by IAT yet missed by FCM were also detected by IAF. This study demonstrates that discordance between IA and FCM is probably not due to cell loss during FCM cell processing or sampling area differences, but may be due to differences in assessing DNA ploidy in the interpretation of IA histograms and/or dilution of aneuploid cells by normal diploid cells in FCM.