Fig. 7. Expression and localisation of Ca²⁺ store ATPases in human spermatozoa. Western blotting was used to detect SERCAs (A) and SPCA1 (B) in rat brain (br) and sperm (sp) lysates. A robust signal, at the appropriate molecular weight, was obtained with the SERCA antibody (Y1F4) in brain lysates but no staining was detected with sperm (three experiments). In contrast, using the same lysates, we detected SPCA1 both in rat brain and in sperm. Though the intensity of the band was considerably lighter in sperm than in brain, the band was found consistently. The blot has two sperm protein lanes to establish that the band was not due to `bleed' from the brain lysate lane. (C-F) In-situ localisation of SERCA and SPCA1 in human spermatozoa. Pictures show overlays of fluorescence and phase-contrast images. All fluorescent images were obtained and processed using identical procedures. Immunolocalisation using antibody Y1F4 showed no significant staining (C) whereas the antibody directed against SPCA1 (D) localised clearly to the rear head and midpiece. Incubation with the secondary antibody alone gave no significant labelling in either case (E and F). Scale bar in C applies to C-F.