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Co-localization IHC experiments are performed to determine the distribution of staining of two different antibodies on the same tissue section. This method allows the investigator to identify whether the same cell is expressing both antigens, and where they are localizing within the cell. The procedure is performed sequentially, by first performing immunochemistry with an antibody using an AP-Vector Red detection system, followed by applying the second antibody using an HRP-DAB detection system. The resulting colorometric precipitates allow for simultaneous detection of both targets on a single section because the first antibody will produce a red deposit, and the second will appear brown. When the antibody co-localizes, the deposit will appear reddish-brown. Prior to performing co-localization IHC, it is important to test each primary antibody individually and determine the antibody staining concentration range that produces the optimal signal for that tissue sample.
The following steps are to be conducted at room temperature. Do not allow tissues to dry at any time during the staining procedure.
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