Muratori Monica, Forti Gianni, Baldi Elisabetta
Andrology Unit, Department of Clinical Physiopathology, Center for Research Transfer and High Education DeNothe, University of Florence, I-50139 Firenze, Italy.
Cytometry A. 2008 Sep;73(9):785-7. doi: 10.1002/cyto.a.20615.
Conflicting results are reported by recent studies comparing flow cytometry (FCM) and fluorescence microscopy (FM) for detecting sperm DNA fragmentation by TUNEL assay. Each of the two technologies has specific peculiarities and limitations, but whereas the limitations of FM observation are well known, the biases due to the inability of FCM to recognize morphologically analyzed cells are less explored. In particular, so far, FCM analysis of sperm DNA fragmentation have included in the analyses M540 bodies, round semen structures exhibiting FSC/SSC properties similar to sperm. Semen M540 bodies, altogether with the occurrence of two sperm populations with different nuclear staining, concur to an underestimation of values of sperm DNA fragmentation by FCM. However, even considering such bias, the observed discrepancies between the performance of FM and FCM are not fully explained. We discuss here the possible variables that may affect the results of each of the two technologies and the necessary efforts to be taken to address this issue.
最近的研究比较了流式细胞术（FCM）和荧光显微镜（FM）通过TUNEL检测精子DNA片段化的结果，结果相互矛盾。这两种技术都有各自的特点和局限性，虽然FM观察的局限性众所周知，但FCM无法识别经形态学分析的细胞所导致的偏差则较少被探讨。特别是，到目前为止，精子DNA片段化的FCM分析在分析中纳入了M540小体，即圆形精液结构，其展现出与精子相似的FSC/SSC特性。精液M540小体，连同出现的具有不同核染色的两个精子群体，共同导致FCM对精子DNA片段化值的低估。然而，即使考虑到这种偏差，FM和FCM性能之间观察到的差异也没有得到充分解释。我们在此讨论可能影响这两种技术各自结果的变量，以及为解决此问题需要付出的努力。
