Assessment of Pisum sativum agglutinin in identifying acrosomal damage in stallion spermatozoa.
The use of fluorescein-conjugated Pisum sativum agglutinin (FITC-PSA) was evaluated for its ability to distinguish acrosome-intact from acrosome-damaged stallion spermatozoa. Incubation of fresh (acrosome-intact) and frozen-thawed (acrosome-damaged) spermatozoa with FITC-PSA resulted in acrosome-intact spermatozoa that exhibited no fluorescence, while acrosome-damaged spermatozoa exhibited fluorescent staining over the rostral portion of the head and equatorial segment. Experiments using mixtures of various ratios of acrosome-intact and acrosome-damaged spermatozoa determined the precision (intrasample coefficient of variation), and linearity (increased percentage of spermatozoa with PSA binding, with increased percentage of frozen-thawed spermatozoa in a sample) of FITC-PSA binding. The binding of FITC-PSA increased in samples as the portion of frozen-thawed (acrosome-damaged) to fresh (acrosome-intact) spermatozoa increased. A positive correlation existed (r = 0.98, P less than 0.05) between the percentage of FITC-PSA bound sperm and the proportion of damaged spermatozoa added to a sample. Location of PSA lectin binding on acrosome-damaged spermatozoa, determined by electron microscopy using gold-conjugated PSA, was to components of the outer acrosomal membrane and acrosomal matrix. These results demonstrate that FITC-PSA binding may be useful in determining acrosomal integrity of fresh and frozen-thawed stallion spermatozoa.
Farlin
M E
ME
Animal Reproduction and Biotechnology Laboratory, Colorado State University, Fort Collins 80523.
Jasko
D J
DJ
Graham
J K
JK
Squires
E L
EL
D000177:Acrosome / Q000648:ultrastructure*; D000818:Animals; D015925:Cryopreservation; D016650:Fluorescein-5-isothiocyanate; D006736:Horses / Q000502:physiology*; D037102:Lectins / Q000378:metabolism; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D037121:Plant Lectins; D012662:Semen Preservation
D037102:Lectins; D008562:Membrane Glycoproteins; D037121:Plant Lectins; C036292:pea lectin; D016650:Fluorescein-5-isothiocyanate