Freshly ejaculated human sperms were washed in PBS and extracted in 3% acetic acid, 10% glycerol, 30 mM benzaminidine. The acid extract was dialyzed against 0.2% acetic acid and subsequently used for immunization.
VCP (valosin-containing protein), also known as p97, TERA, ALS14, IBMPFD, HEL-220, IBMPFD1, or HEL-S-70, is a member of a protein family that includes putative ATP-binding proteins involved in vesicle transport and fusion, 26S proteasome function, and assembly of peroxisomes. VCP is a structural protein that associates with clathrin and heat-shock protein Hsc70, to form a complex. It has been implicated in a number of cellular events that are regulated during mitosis, including homotypic membrane fusion, spindle pole body function, and ubiquitin-dependent protein degradation. In sperm this intra-acrosomal protein can be used as a marker for evaluation of the physiological state of sperm cells as well as for selection of a suitable method of fertilization in the laboratories of assisted reproduction. Applications: Suitable for use in Western Blot, Flow Cytometry and Immunocytochemistry. Other applications not tested. Recommended Dilution: Flow Cytometry: 3-12ug/ml, Intraacrosomal staining Immunocytochemistry: 10ug/ml, membrane permeabilization (acetone) is essential. Designed for quantitative immunofluorescence analysis of pathological sperms (excellent tool for laboratories of assisted reproduction when optimal method of fertilization is sought). Optimal dilutions to be determined by the researcher. Storage and Stability: May be stored at 4C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20C. Aliquots are stable for 12 months after receipt. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
