Abstract
We investigated the optimal gold particle size for use with polarized incident light (epi polarization) microscopy with immunogold immunohistological preparation in both immunogold indirect (IGS) and silver-enhanced immunogold-silver staining (IGSS) techniques. A range of gold particle sizes from 5 nm-40 nm was used along with tissue of known immunoreactivity with a well-characterized primary monoclonal antibody. Checkerboard titrations were carried out for each technique and for each particle size. The preparations were viewed using a standard polarized incident light microscope and assessed in a semi-quantitative manner. Adequate visualization of gold particles was achieved using the indirect staining method only with a particle size of 40 nm. With silver enhancement (IGSS), particles of all sizes were clearly seen. However, 5-nm particles were considered optimal for this method because of reduced background staining, high titration of antisera possible, and crisp localization of the visual signal.