Quantitative analysis of post-embedding immunogold Electron Microscopy with differently sized labelling gold particles

Abstract

The post-embedding immunogold staining technique is a powerful tool in localizing tissue antigens at the ultrastructural level. The highly electron dense colloidal gold particles are easily visible under electron microscope and the differently sized gold particles allow a simultaneous demonstration of more than one tissue antigens in the same grid. The sizes of the commercially available labelling gold particles range from 1 nm to 40 nm. It is generally believed that the smaller the gold particles the better the detecting sensitivity of the technique. However, no quantitative data in this regard is available. We carried out an experiment to assess the detecting sensitivities of differently sized labelling gold particles in the post-embedding electron microscopic indirect immunogold staining method using atrial natriuretic peptide (ANP) in rat atria as a model.ANP is a newly discovered cardiac hormone with 28 amino acids. In the heart it is located exclusively in the specific electron dense granules in the cytoplasm of atrial cardiocytes. The high concentration and the well defined location of this tissue antigen makes it an ideal model to evaluate the detecting sensitivities of immunostaining techniques.