Biochemical Techniques and Instrumentation

Materials

1. Bicarbonate/carbonate coating buffer (100 mM)

Antigen or antibody should be diluted in coating buffer to immobilize them to the wells:

Na₂CO₃ - 3.03 g

NaHCO₃ - 6.0 g

Distilled water - 1000 ml

pH 9.6,

2. Phosphate buffered saline (PBS)

Na₂HPO₄ - 1.16 g

KCl - 0.1 g

K₃PO₄ - 0.1 g

NaCl - 4.0 g

Distilled water - 500 ml

pH 7.4.

3. Blocking solution

Commonly used blocking agents are 1% BSA or 5% non-fat dry milk in PBS.

4. Wash solution

Usually PBS or Tris -buffered saline (pH 7.4) with detergent such as 0.05% (v/v) Tween20.

5. Antibody dilution buffer

Primary and secondary antibody should be diluted in 1X blocking solution to reduce non specific binding.

6. Substrate

7. Enzyme conjugale

8. Stop solution