Characterization of antigenic proteins in dog spermatozoa and effect of immunization with sperm membrane proteins on semen quality

In this paper, the use of hetero- and iso-antibodies to identify and characterize sperm-specific antigenic proteins in dogs is applied. Also, the effect of antisperm antibodies on semen quality is represented. Dog sperm proteins extracted with sodium dodecyl sulphate (SDS-SME) and lithium diiodosalicylate (LDIS-SME) were injected into one male dog each and SDS-SME were also injected into 2 male rabbits. A rapid humoral response was confirmed from a strong precipitation line in immunodiffusion and an increase in immunoglobulin (Ig) level in the blood serum. A maximum titre of 1:3200 was achieved between 77 days of post immunization (DPI) to 117 DPI and on 35 DPI in dog and rabbit blood serum, respectively. Partially purified iso-Ig of dog detected nine antigenic proteins of 97, 46, 42, 38, 36, 32, 30, 28 and 20 kDa in dog sperm whereas, partially purified hetero-Ig of rabbits recognized only three protein bands of 97, 38 and 36 kDa in dog sperm. The cross reaction of anti LDIS-SME and anti SDS-SME antibodies with 46 and 28 kDa protein bands in epididymal fluid, tissue and blood plasma, indicated the involvement of these antigens in sperm maturation during epididymal transit and their circulation in blood. Fourteen and 116 kDa antigens, recognized by anti-SME in blood serum/tissue/fluid, might be non-specific proteins. On using fluorescein isothiocyanate (FITC) labeling on dog spermatozoa with rabbit Ig, it was observed that antigens recognized by the Ig were located at the surface of the spermatozoa. Localization of antigenic proteins was more intense on the acrosomal cap. A significant decrease (p<0.05) in motility, live spermatozoa, HOST, sperm count and an increase in sperm abnormalities was observed in the ejaculates of two dogs from 45–90 and 90–150 DPI. An improvement in sperm parameters from 150 DPI onwards was observed, which reverted equivalent to that of pre-immunized dog ejaculates after 210 DPI. Therefore, it can be concluded that dog SME has immuno-contraceptive potential and in order to achieve a better characterization of these dog antigenic proteins, and before understanding their role in the dog-gamete interaction or considering their use in a contraceptive vaccine, their molecular structure, specificity and function need to be investigated.