Becker R L, Mikel U V
Department of Cellular Pathology, Armed Forces Institute of Pathology, Washington, D.C. 20306-6000.
Anal Quant Cytol Histol. 1990 Oct;12(5):333-41.
The severity and consistency of the effect of formalin fixation on the quantitation of DNA by flow cytometry (FCM) and image cytometry (ICM) were studied. As compared to ethanol, formalin fixation substantially decreased the propidium iodide fluorescence from mouse hepatocyte nuclei analyzed by FCM; it was also associated with an altered 4n-to-2n signal ratio and with false aneuploid peaks by FCM, but not by ICM (microspectrophotometry). ICM, on the other hand, suffered from a dependence of the DNA signal on nuclear size, which was not seen with FCM. The DNA signal variation was related to variations in the chromatin state, as shown by differences between monocytes and lymphocytes, and between RAJI cells fixed under various ionic strengths. The dependence of the DNA signal on the chromatin state indicates a need for caution in interpreting aneuploidy in formalin-fixed cells. For FCM, pseudoaneuploidy appears avoidable by using a Feulgen fluorescence staining technique. New imaging modes may be necessary to solve the problem of cell size dependence for ICM DNA determination.
研究了福尔马林固定对通过流式细胞术（FCM）和图像细胞术（ICM）进行DNA定量的影响的严重程度和一致性。与乙醇相比，福尔马林固定显著降低了通过FCM分析的小鼠肝细胞核的碘化丙啶荧光；它还与FCM中改变的4n与2n信号比率以及假非整倍体峰相关，但ICM（显微分光光度法）则没有。另一方面，ICM存在DNA信号对核大小的依赖性，而FCM则没有这种情况。DNA信号变化与染色质状态的变化有关，如单核细胞和淋巴细胞之间以及在不同离子强度下固定的RAJI细胞之间的差异所示。DNA信号对染色质状态的依赖性表明在解释福尔马林固定细胞中的非整倍体时需要谨慎。对于FCM，通过使用Feulgen荧光染色技术似乎可以避免假非整倍体。可能需要新的成像模式来解决ICM DNA测定中细胞大小依赖性的问题。
