Immunolocalization of secretory protein-I or chromogranin A in amphibian urinary bladder granular cell granules

The presence of secretory protein-I (SP-I) or chromogranin A (CGA) in granules isolated from the granular cells of the amphibian urinary bladder epithelium was investigated using ultraimmunohistochemistry. Granules were isolated by cell fractionation using Percoll density gradients. SP-I was isolated and purified from bovine parathyroid glands. Antibodies were raised in rabbits and purified by affinity chromatography. Ultraimmunohistochemistry, employing the avidin-biotin-peroxidase (ABC-complex) procedure, was used to localize SP-I on thin sections of isolated granules. About 27% of the granules from control (−ADH) cells were SP-I+, while 51% of the granules fractionated from hormone treated (+ADH) cells were positive for this protein (p < 0.0001). Accordingly, granules from ADH-treated cells also showed a significant (p < 0.0001) increase in total protein.