Antibodies are initially titered by spectrophotometry using a NanoDrop spectrophotometer. If needed, the sample is adjusted by diluting or concentrating to a final concentration of 0.9-1.1 mg/mL. Antibody concentration is confirmed by Coomassie staining.
For Coomassie staining, the sample is separated via a denaturing SDS-PAGE gel alongside serial dilutions of a standard antibody of known concentration and stained with Coomassie blue. Protein band intensities are measured using ImageJ software and a standard curve is generated. The final antibody concentration is extrapolated from the standard curve.
Using ImageJ software, the band intensities of both the antibody and impurities are measured and the ratio of the antibody protein content to the total protein content of the sample is determined. Only antibody samples with >90% purity are distributed.