Assessment of Pisum sativum agglutinin in identifying acrosomal damage in stallion spermatozoa

Abstract 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 (acrosomeintact) 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 frozenthawed 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 (acrosomeintact) spermatozoa increased. A positive correlation existed (r = 0.98, P < 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. © 1992 Wiley‐Liss, Inc.