Immunocytochemical assays of amylase and chymotrypsinogen in rat pancreas secretory granules. Efficacy of using immunogold-labeled ultrathin cryosections to estimate relative protein concentrations

Abstract

An exploration was conducted as to whether the relative concentration of two intracellular proteins could be evaluated quantitatively from their labeling densities in ultrathin cryosections labeled with the immunogold technique. As a model rat pancreatic cells were used in which the content of amylase (Am) and chymotrypsinogen (Ch) was experimentally altered. Rats were fed either normal laboratory chow or food containing soybean trypsin inhibitor (STI), which affects the Am/Ch ratio in the tissues. The changes in Am and Ch protein levels and enzyme activities were measured biochemically in cell suspension homogenates or in zymogen granule fractions. Within 5 days a maximal change in the Am/Ch was observed as a result of adaptation to the STI diet. The Am/Ch ratio determined biochemically was compared with that from counts of gold particles bound to the respective protein in immunogold-labeled cryosections. The two data sets matched fairly well, indicating that the intensity of the immunoreaction is a reliable reflection of antigen concentration in this system.