High Density Protein Arrays for Antibody Validation

IN ONCYTE AVID, ONCYTE nitrocellulose film slides, Product Applications, Uncategorized

A recent paper in BMC Biotechnology by D. Ma et. al of Origene describes the use of high density protein arrays to test the specificity, or lack thereof, for antibodies used for immunoassay diagnostics. The antibody 8F1 is  commonly used in immunohistochemistry assays to detect ERCC1 protein, a biomarker for testing the efficacy of cisplatin-based chemotherapy used in non-small lung cancer therapy.Screening with a high density protein array demonstrated the cross-reactivity of the 8F1 antibody with a nuclear protein, thus clouding its use as a diagnostic tool. Further, two other antibodies to ERCC1 showed no cross-reactivity with other proteins, and demonstrate higher specificity than the 8F1 antibody. To generate the high density array of 10,464 proteins the group from Origene used their TrueORF DNA transfection system in HEK293T cells, then spotted  cell lysates onto AVID nitrocellulose film slides from Grace Bio-LabsOrigene currently offers a variety of tissue lysate arrays as well as cell lysate  arrays for cancer research. While the array used in this study represented a small fraction of the anticipated human proteome, it did demonstrate utility in discriminating relative specificity among several antibodies. Most importantly, the 8F1 antibody, while the most frequently cited tool for detection of ERCC1, may be retired from the researcher’s toolbox. The high density array described in the paper is not yet available on the Origene website, but may prove a useful tool in future to discriminate antibody specificity and selection of the best tools for immunoassays.