Add Secondary Antibody Conjugated To Alkaline Phosphatase

FIGURE 26.06 Blot

Western

After a mixture of proteins have adhered to the nitrocellulose membrane, one specific protein can be detected using an antibody. The antibody is added with a solution of milk proteins and incubated with the nitrocellulose membrane. The milk proteins bind to and block those regions of the nitrocellulose that do not have any protein attached. The primary antibody attaches only to the protein of interest. A secondary antibody with alkaline phosphatase attached binds specifically to the primary antibody, and allows the single protein band to be visualized.

Secondary antibody binds to primary antibody

Secondary antibody binds to primary antibody

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