ABSTRACT
One of the most widely used and sensitive of the qualitative immunological techniques is that of immunoblotting, or Western blotting. When combined with the technique of sodium dodecyl-sulfate polyacrylamide gel electrophoresis, immunoblotting forms an invaluable means of identifying antibody-antigen reactions within complex mixtures of bacterial components. This technique involves the electrophoretic transfer of separated bacterial components from polyacrylamide gels onto sheets of nitrocellulose. For reaction with antibodies, unfilled protein/lipopolysaccharide (LPS) binding sites on nitrocellulose sheets are saturated with a suitable protein, such as casein in skimmed milk powder. Immunoblotting can also be used to screen antisera for specific antibodies by reacting sera with well-characterized proteins or carbohydrates. When analyzing human sera always assume that the patient was infected with human immunodeficiency virus or hepatitis B virus and take the necessary precautions. The chapter describes the results of two applications of the immunoblotting technique.
