For recombinant protein immunoblotting, selected mAbs were used to probe 50, 10, 2, and 0.4 ng of recombinant protein antigens. For immunoblotting of endogenous human protein samples by PETAL mAbs, cell lines were selected according to the protein expression profile from HPA and UniProt databases. For membrane or nuclear proteins, corresponding cellular fractions were prepared for immunoblotting. Typically, 20 μg of protein was loaded onto each lane. Support-positive immunoblotting results were evaluated following the criteria described by Antibodypedia ( and HPA (29). Basically, an antibody was qualified as immunoblotting positive when the size of a single or predominant single band on immunoblotting matched or was within 10% of the predicted antigen molecular weight. In some cases, an immunoblotting-positive conclusion was enhanced when the same predicted protein band was detected in two or more different cell lysates. Some antibodies detected multiple bands with different sizes, but the predicted size protein band was also detected.

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