How much protein amount should be loaded on the gel?

This depends on several factors:

(1)  The sample type

For total protein extracted from cell or tussle lysate, 10-40 μg is recommended; for purified protein, 10-100 ng is recommended. A compatible protein assay (e.g., BCA Assay, Bradford Assay, etc.) to determine the protein concentration of each sample is optional before electrophoresis, as a way to avoid under- or over-loading of samples.

(2)  The abundance of the target protein in the lysate

(3)  The gel type

The numbers of wells in a gel also determine the amount of each sample loading.

(4)  The affinity of the antibody to a specific protein

It should be noted not to overload each well in the gel, otherwise, the bands get fused.

Hello Dalal,

From my experience, it's better to test load various amounts as you want to put enough of your protein of interest it's usually a balance between putting enough not too little but not too much and your Ab that you are using plays a huge role. it would be best if you optimised, in your case you should aim between 10 to 25 ug per well as I suppose you are not overexpressing and purifying.