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Comparing flow cytometry and fluorescence microscopy for analyzing human sperm DNA fragmentation by TUNEL labeling
Author(s) -
Muratori Monica,
Forti Gianni,
Baldi Elisabetta
Publication year - 2008
Publication title -
cytometry part a
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.316
H-Index - 90
eISSN - 1552-4930
pISSN - 1552-4922
DOI - 10.1002/cyto.a.20615
Subject(s) - dna fragmentation , sperm , flow cytometry , fragmentation (computing) , biology , semen , fluorescence microscope , staining , tunel assay , dna , microbiology and biotechnology , fluorescence , genetics , apoptosis , physics , optics , ecology , programmed cell death
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. © 2008 International Society for Advancement of Cytometry