Western blotting is used to indentify or detect any specific protein in a given sample. In this technique, first protein molecules are gel electrophoresed with polyacrylamide gel electrophoresis and transferred to nitrocellulose or nylon membrane. Then immobilized protein is detected by specific antibody which binds only to the protein of interest at specific temperature and pH, this antibody is referred as primary antibody. Primary antibody could be labeled radioactively or may be tagged to radioactive molecule to directly detect the protein. If the primary antibody is not labeled, in this case another antibody is used which has binding site for primary antibody and is radioactive or linked or conjugated with enzyme or fluorescent tags i.e. labeled to make the presence of protein visible to eye. This second antibody which binds to primary antibody is referred as secondary antibody
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Western Blotting
