Small colloidal gold conjugated to Fab fragments or to immunoglobulin G as high-resolution labels for electron...

Fab-colloidal gold labelling in conjunction with negative staining and high-resolution electron microscopy was used for targeting single protein units in regular arrays. These were bacteriophage T4 polyheads with Fab-Au2.5, and a specific antibody binding site on the haemagglutinin polypeptide of influenza virus with Fab-Au3, Fab-Au2.5, and Fab-Au1-2. For the latter, IgG-Au3 was also used. Experimental details are summarized to provide generally applicable methods for the preparation of small gold colloids Fab-Au and of labelling. The putative mechanism of protein-gold complex formation and adsorption to preferred sites on Fab and IgG, most probably to sulphur-rich regions, is discussed. The influence of pH during complex formation was found to be of minor importance in the samples investigated. Reported experimental details and our own experiences suggest that the importance of a protein's pI relative to its optimum gold complexing pH critically depends on the nature of the protein in question rather than being of general importance for protein-gold complex stability.