RESIN EMBEDDING FOR QUANTITATIVE IMMUNOELECTRON MICROSCOPY - A COMPARATIVE COMPUTERIZED IMAGE-ANALYSIS

Quantitative immunoelectronmicroscopy (QIEM) is dependent on the relia bility of preparative techniques for both efficient immunolabeling and consistent quantitative results among series of immunostained section s. The present study compared Lowicryl K4M and Epon embedding after id entical fixation and dehydration of rat somatotrophic secretory granul es. Labeling intensity, diameter, roundness, uptake of uranyl acetate, and gray value were measured with computer assisted image analysis. L owicryl-embedded granules showed the highest labeling densities after conventional fixation and Progressively Lowering Temperature (PLT) deh ydration, but values were not consistent in a series of immunostained sections. A lower but more uniform level of immunostaining was seen in Epon-embedded sections when tissue was cryofixed and physically dehyd rated. Gray value measurements from micrographs from both embedding me dia confirmed the better contrast of Epon sections and the different r eliefs of the granule surfaces. This study emphasizes the importance o f complete comparisons of preparative techniques for QIEM for reliabil ity and reproducibility.