In summary, the 8F1 clone is not suitable for ERCC1 IHC assay due to its cross-reactivity with PCYT1A protein. Two clones showed high mono-specificity on the protein microarray chip test and both worked for the IHC application. To develop the best monoclonal antibody for ERCC1 IHC analysis, 18 monoclonal antibodies were generated and 6 of them were screened against our protein microarray chip. In this study, we also discovered that the PCYT1A gene expression level is significantly higher than the ERCC1 gene expression level in a certain population of lung cancer patient tissue samples. Similar to the subcellular localization of ERCC1, IHC tests demonstrated that PCYT1A is localized mainly on nuclear membrane. The cross-reactivity is due to a common epitope presented on these two unrelated proteins. Resultsīy using a high density protein microarray chip technology, we discovered that 8F1 not only reacts with its authentic target, ERCC1, but also cross-reacts with an unrelated nuclear membrane protein, PCYT1A. 8F1 is one of the most commonly used monoclonal antibodies for evaluating ERCC1 expression levels in lung cancer patient tissues, but it has been noted that this antibody cross-reacts with an unknown protein. High ERCC1 expression is linked to drug resistance on cisplatin-based chemotherapy. ERCC1 is being explored as a predictive diagnostic biomarker for cisplatin-based chemotherapy. An antibody with cross-reactivity can create unexpected side effects or false diagnostic reports if used for clinical purposes.
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