1. Dot blot can be used to estimate protein concentration in crude or partially purified samples.

2. Cut nitrocellulose membrane to the appropriate size to fit in the dot-blot apparatus. Wet the membrane by immersing it in deionized water. For PVDF membrane pre-wet with methanol for seconds and soak in deionized water. Add sample and standard diluted in 100 m l of PBS/0.05% Tween 20 to each well. Draw the liquid through the membrane by applying suction pump.

2a. Alternatively cut nitrocellulose membrane (not PVDV) to the appropriate size. Draw grids with a pencil. Spot 1 m l of the sample and standard in the center of the grid. Let the membrane dry.

You may make serial dilution of the sample and standard.

3. Proceed to Western.