This Immuno-Fixation is followed by the ammoniacal silver staining. Immuno-fixation and silver staining simply done inside the gel. The silver staining starts from step 5, no TCA-fixing and washing necessary. This method combines the sensitivity of the silver-staining with the specificity of the antibody-reaction.
Advantage: Only IgG will be stained, band-sharpness and sensitivity as good as with the general silver-staining. Disadvantage: One overnight step (washing out of the anti-human IgG serum).
The Immunoreaction is done in the CleanPool in 30 ml (100 µl Antibody), 2 hours slightly rocked.
Overnight: Washing out exceed amount of the antibody
Silver staining can be automated with GE's Autostainer
Gel: FocusGel 6-11 24S Visualization: Immuno-Fixation followed by silver staining (Courtesy of G.Cox, Charlton Memorial Hospital, Swansea)