The blocking buffer should improve the sensitivity of the assay by reducing background interference. Individual blocking buffers are not compatible with every system. For this reason, a variety of blockers in both Tris-buffered saline (TBS) and phosphate-buffered saline (PBS) are available. The proper choice of blocker for a given blot depends on the antigen and on the type of enzyme conjugate to be used. For example, with applications using an alkaline phosphatase conjugate, a blocking buffer in TBS should be selected because PBS interferes with alkaline phosphatase. The ideal blocking buffer will bind to all potential sites of nonspecific interaction, eliminating background without altering or obscuring the epitope for antibody binding.
