Western Blot - 6. Buffers and Blocking Reagents

When performing a western blot, the blocking buffer should not be overlooked. The blocking buffer fills in the locations on the membrane that can still bind protein and cause background if not treated. The critical reagent in a blocking buffer is protein, where the protein is non-antibody reactive. Popular blocking proteins include non-fat dried milk (NFDM), BSA, casein, and combinations thereof. Of note, a single blocking agent may not be sufficient for all western applications. Some blocking agents can interfere with primary antibody activity, or may not be compatible with the reporter system in use, or produce undesired auto-fluorescence. Rockland is able several blocking buffer reagents suitable for all western blot applications, including BLOTTO-NFDM and BSA for standard applications, and a specially formulated Blocking Buffer for Fluorescent Western Blotting.

Western Blotting Protocol

1. Background
2. Types of Western Blot Formats
3. Primary Antibody Reagents
4. Secondary Reporter Antibody Reagents
5. Substrates
6. Buffers and Blocking Reagents
7. Western Blot Preassembled Kits