Factors affecting the consecutive and simultaneous coupling with bis-diazotized benzidine of protein antigen to sheep red blood cells

The dynamics of attaching antigen to sheep red blood cells using bis-diazotized benzidine (BDB) were studied using purified protein derivative tuberculin as antigen. Factors considered were the concentration of antigen and BDB, temperature, time and simultaneous or consecutive coupling. Simultaneous coupling at 42°C for 10 min gave the most specific haemagglutination reactions and was simple to carry out.