Acrosome Biogenesis in the Hamster: RESULTS(6)
AM22/AM29 Expression during Spermatogenesis
Light and electron microscopic immunolabeling was employed to define the temporal expression and localization of the acrosomal matrix polypeptides in spermatogenic cells. When polyclonal anti-AM22 was used, protein expression was noted in the acrosome at all stages of spermatid development (Fig. 9, A, B, D, and E). In late matu-ration-phase spermatids, the staining with anti-AM22 became restricted to the apical and principal segments of the acrosome whereas the equatorial segment was negative (long arrows on Fig. 9, D and E). When monoclonal anti-AM29/22 was used, the acrosome of late maturation-stage spermatids exhibited intense fluorescence staining (long arrows Fig. 9, С and F); however, the acrosome of earlier spermatids was negative (Fig. 9, С and F). Ultra-structural immunolabeling using anti-AM22 demonstrated labeling throughout the matrix of the acrosomal vesicle of round spermatids (Fig. 10). These data indicate that AM29 and/or AM22, or alternatively a precursor protein, is expressed in early spermatids. Moreover, the data suggest that an intra-acrosomal posttranslational protein-processing event, occurring in late maturation-phase spermatids, is required for antigen recognition by monoclonal anti-AM29/ 22. buy diabetes drugs
FIG. 9. Matched phase contrast (A, D) and fluorescence (В, С, E, F) images of spermatogenic cells that were doublestained with polyclonal anti-AM22 and monoclonal anti-AM29/22. В and E used filter sets to visualize staining with polyclonal anti-AM22 while С and F used filters to visualize staining with monoclonal anti-AM29/22. Note that round spermatids (rs) show fluorescent acrosomes (a) when immunostained with polyclonal anti-AM22 (В, E) but not when stained with monoclonal anti-AM29/22 (C, F). Late maturation-phase spermatids (long arrows) show intense acrosomal fluorescence with both polyclonal (В, E) and monoclonal (C, F) antibody, whereas early maturation-phase spermatids (short arrow) stain only with polyclonal antibody (E, F). Bar = 10 (j.m.