title: Comparison of rat epididymal sperm counts by IVOS HTM-IDENT and hemacytometer.
abstract: Epididymal sperm counts, a common measurement in male reproductive toxicity studies, are routinely determined using a hemacytometer. Recently, computer assisted methods for automated sperm counts have been developed. In the present study we evaluated an automated system, the TOX IVOS (Hamilton Thorne Research, Beverly, MA) HTM-IDENT option, that utilizes a DNA-specific stain and fluorescence illumination to identify sperm for enumeration. Cauda and caput epididymal sperm counts were determined in 48 adult male Sprague-Dawley rats, using both the hemacytometer and HTM-IDENT. The mean hemacytometer and HTM-IDENT counts (+/- SD) were 250 +/- 43 and 254 +/- 52 million, respectively, for cauda sperm, and 123 +/- 13 and 127 +/- 18 million, respectively, for caput sperm. The average coefficient of variation using the hemacytometer was 13.8% as compared to 17.3% for the HTM-IDENT. Comparison of the machine count and a visual count from the Display Statics screen of the HTM-IDENT indicated that when two or more sperm heads touched or overlapped, the machine counted them as one. Manual (visual) and machine counts when compared over a range of nine concentrations from 3.7 to 47.8 million/mL differed by 4 to 12% at the lowest to highest concentration. The concentration of epididymal sperm samples used in comparing the two counting methods ranged from 5.8 to 17.7 million/mL. Therefore, the HTM-IDENT undercounting error attributable to sperm heads touching was less than 6%. Overall the data indicate good agreement between the HTM-IDENT and the hemacytometer counts. Furthermore, both counting time and technician fatigue were markedly reduced. Thus the HTM-IDENT option improves the efficiency of epididymal sperm counting without loss of precision.
authors.affiliation: Reproductive Toxicology Division, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina 27711, USA.
authors.forename: L F
authors.initials: LF
authors.lastname: Strader
authors.forename: R E
authors.initials: RE
authors.lastname: Linder
authors.forename: S D
authors.initials: SD
authors.lastname: Perreault
mesh_terms: D000818:Animals; D004822:Epididymis / Q000166:cytology*; D007091:Image Processing, Computer-Assisted; D008297:Male; D008853:Microscopy; D051381:Rats; D017207:Rats, Sprague-Dawley; D013076:Sperm Count / Q000379:methods*