✍🏼 Xiaofeng Li, Qiana Mendez, Cassandra Chapados, Felicity Acca, Holly Driscoll, Jason Oliveira, Jun Liu, Kezzia Jones, Mary Ferguson, Ryan L. Wallace, Sergei Bibikov, Troy Lionberger, Kevin J. Harvey, Michael P. Weiner, Greg Mirando
🏠 Abbratech, Branford, CT, USA
📑 New Biotechnology (2025)
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Abstract
Kirsten rat sarcoma viral oncogene homolog (KRAS) is the most mutated oncogene in human cancers, found in approximately 30 % of tumors. These mutations primarily consist of single-base missense alterations in codon G12. While extensive efforts have focused on developing allele-specific inhibitors for KRAS mutations, mutation-specific antibodies (Abs) remain largely unexplored, with only a few research-use-only catalog Abs available. In this study, we employed the proprietary Epivolve technology to develop site-directed monoclonal Abs (mAbs) that target KRAS oncogenic driver mutation KRAS G12D. These site-directed mAbs demonstrate high binding affinity, with equilibrium dissociation constants (KD) in the nanomolar range, showing over 1,000-fold greater affinity for KRAS G12D compared to wild-type KRAS. Western blot analyses using both purified KRAS protein variants and tumor cell lines harboring G12D mutations confirmed the high specificity of these mAbs. Furthermore, immunocytochemistry analysis revealed co-localization of the site-directed mAbs with endogenously expressed KRAS in cancer cells bearing G12D mutations. The validated high affinity and specificity of these site-directed mAbs highlight their potential for diagnostic applications and therapeutic development targeting KRAS driver mutations.
How the WOLF was used in this study
In the New Biotechnology article (S187167842500041X), the authors utilized the WOLF Cell Sorter (NanoCellect Biomedical) during their flow cytometry workflow to isolate specific immune cell populations, including B cells, by fluorescence-activated sorting. After staining cells with viability and fluorescence markers such as SYTOX to discriminate live from dead cells, the researchers ran the suspension on the WOLF sorter to enrich for live, targeted cell populations with defined surface phenotypes. This use of the WOLF instrument helped ensure that only viable, phenotypically correct cells were collected for further downstream analysis and characterization in their antibody discovery and binding studies.
