Radioimmunoassays for Bull Seminal Plasma Proteins (BSP-A1/-A2, BSP-A3, and BSP-30-Kilodaltons), and Their Quantification in Seminal Plasma and Sperm1

Abstract

Three proteins, BSP-A1/-A2, BSP-A3, and BSP-30 kilodaltons (collectively called BSP proteins), represent the major proteins of bovine seminal plasma (BSP). At ejaculation, these proteins bind to the sperm surface and induce molecular changes in the plasma membrane that are deemed to be essential for sperm capacitation. The present study was carried out to develop specific radioimmunoassays (RIAs) for the quantification of each of the BSP proteins in BSP and sperm. RIAs were developed using polyclonal antibodies raised in rabbits against each BSP protein. The purified and iodinated BSP proteins were used as standard and tracer, respectively. The RIAs that were developed were shown to be specific for each protein and the cross-reactivity toward various antigens was negligible (<2%). The average sensitivity limit was 5 ng/ml of sample for BSP-A1/-A2 and BSP-A3, and 40 ng/ml of sample for BSP-30-kDa. The concentration of BSP proteins was determined by analyzing the RIA data with spline function. BSP proteins represented 40% to 57% of seminal plasma total protein (25% to 47% of BSP-A1/-A2, 3% to 5% of BSP-A3, and 3% to 7% of BSP-30 kDa) and 4% to 6% of sperm total protein (2.5% to 4% of BSP-A1/-A2, 0.4% to 0.9% of BSP-A3, and 0.5% to 1% of BSP-30-kDa). We also determined the concentration of BSP proteins that were sperm-bound after semen cryopreservation in Tris-egg yolk-glycerol extender. A significant decrease (70%-80%) in sperm-bound BSP proteins was noted after cryopreservation. The availability of reliable RIA procedures should aid in the further understanding of the role of BSP proteins in sperm function as well as their effect on sperm cryopreservation.

Seminal Plasma Proteins as Potential Markers of Relative Fertility in Zebu Bulls (Bos taurus indicus)

“…The concentration of BSP-A1/-A2 was much higher compared with other seminal plasma proteins, and this protein alone represented a average of 38% of the total protein fraction, whereas BSP-A3 and BSP-30 kDa represented 3% to 4% of the total protein fraction (Nauc et al, 2000). The determination of BSP protein content on sperm surface may be an index of individual bull fertilizing ability or post-thaw status of sperm membranes (Nauc et al, 2000).…”

“…In the context of sperm cryopreservation may lead to a decrease in sperm resistance to cold shock (Darin-Bennett & White, 1977;White, 1993). Thus, changes induced by the BSP proteins in the sperm membrane may have influence on sperm fertilizing ability and the success of the cryopreservation process (Nauc & Manjunath, 2000).…”

“…The determination of BSP protein content on sperm surface may be an index of individual bull fertilizing ability or post-thaw status of sperm membranes (Nauc et al, 2000).…”

Improving bovine semen diluents: insights from the male and female reproductive tracts, and the potential relevance of cervical mucins

“…BSP proteins are the most abundant proteins in seminal plasma, accounting for 40% to 57% of the total protein present within seminal plasma (Nauc and Manjunath, 2000). Several members of the BSP family of proteins have been described (Nauc and Manjunath, 2000).…”

“…Several members of the BSP family of proteins have been described (Nauc and Manjunath, 2000). Interaction between bovine sperm and BSP1, BSP3 and BSP5 seems to be important for sperm binding to the oviduct as they confer on epididymal sperm increased binding affinity for oviduct epithelial cells (OECs) (Gwathmey et al, 2006).…”