The performance of the assay will depend greatly on the abundance of the protein of interest in the particular cell type or tissue type to be studied. The assays seldom benefit from more than 20 µg of lysate in the well. The minimum amount that can be used will depend on the abundance of the phosphoprotein and the percentage that is phosphorylated within the sample of interest. The most robust assays can be performed with less than 1 µg of total protein.