Comparison between propidium iodide and 7-amino-actinomycin-D for viability assessment during flow cytometric analyses of the human sperm acrosome.
Evaluation of the acrosome reaction can shed light on the fertilising competence of spermatozoa. To eliminate false-positive results when evaluating the acrosome status of human sperm cells, two viability probes propidium iodide (PI) and 7-amino-actinomycin D (7-AAD) were compared for their ability to stain nonviable cells post-fixation and permeabilisation. Both the mean fluorescence and % dead cells differed significantly with time (P < 0.0001). Unlike PI, 7-AAD did not leach from cells and fluorescence remained stable for up to 4 h. Furthermore, 7-AAD proved to be a proficient marker to exclude dead sperm cells during flow cytometric evaluation of ionophore-induced acrosome reaction.
Falzone
N
N
Department of Biomedical Sciences, Tshwane University of Technology, Pretoria, South Africa. falzonen@tut.ac.za
Huyser
C
C
Franken
D R
DR
D000177:Acrosome / Q000378:metabolism / Q000502:physiology*; D020101:Acrosome Reaction; D000328:Adult; D003609:Dactinomycin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D005189:False Positive Reactions; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008297:Male; D008856:Microscopy, Fluorescence; D011419:Propidium / Q000737:chemistry / Q000378:metabolism*; D013094:Spermatozoa / Q000378:metabolism / Q000502:physiology
D005456:Fluorescent Dyes; D003609:Dactinomycin; D011419:Propidium; C025942:7-aminoactinomycin D