Abstract 1325: Novel recombinant rabbit monoclonal antibodies for cancer biology research
Authors: Ball Alexander , Hsieh Yung Lin , Huang Chun Kai , Huang Sega , Hsieh Ping Kuan , Wu Shang Ru , Chang Yue Yun , Liu Jiming , Peng Po Shin , Lin Chia Yi 
Published: April 2023

Abstract
Recombinant monoclonal antibodies (rAbs) are superior tools for cancer biology research as they offer distinct advantages over standard polyclonal and hybridoma-generated monoclonal antibody reagents. Recombinant technology produces antibodies that can be defined at the primary sequence level and are characterized by consistent performance and supply. These rAb features are in stark contrast to traditional polyclonal and hybridoma-generated monoclonal antibodies, whose lack of dependability has been widely documented as a primary reason for data irreproducibility plaguing biomedical research. In an effort to respond to this challenge, GeneTex has established a recombinant antibody production platform combined with more rigorous validation protocols. The goal is to develop reliable recombinant rabbit monoclonal antibodies for reproducible performance in various experimental applications utilized routinely by cancer biologists. GeneTex’s rAb production protocol is based on the approach described by Starkie et al. (2016). This involves a multi-parameter fluorescence-activated single cell sorting (FACS)-based methodology to identify and isolate antigen-specific IgG+ memory B cells obtained from an immunized rabbit. The heavy and light chain variable region genes from selected cells are PCR-amplified and cloned into plasmids to produce full-length heavy and light chains for a single IgG. These constructs are subsequently introduced together into mammalian cells for expression, meaning that natural pairing of the chains is maintained. This manufacturing strategy allows application-specific testing (e.g., for western blot (WB), immunohistochemistry (IHC), and immunocytochemistry (ICC/IF), etc.) of clones during the antibody production process. Validation of antibodies is performed inhouse and, when possible, in relationships with academic or industry/pharma institutions where ideal samples and reagents are frequently more available. This recombinant monoclonal antibody production platform has created a series of well-validated antibodies against many important targets for cancer biology research. This includes reagents to study tumor immunobiology (e.g., PD-L1), hormone receptors (e.g., ER alpha and androgen receptor variants like ARV7), oncogenic RAS protein mutants (e.g., RAS G12D), transcription factors (e.g., NRF2), and epithelial-mesenchymal transition (EMT) (e.g., E-cadherin), among many others. Extensive validation for multiple applications was performed, with knockdown or CRISPR-based knockout cell lysates employed when possible to establish specificity. In summary, GeneTex is utilizing a FACS-based recombinant rabbit monoclonal antibody production approach to generate and thoroughly validate antibody reagents that will hopefully facilitate reproducible cancer biology research.
Citation Format: Alexander Ball, Yung Lin Hsieh, Chun Kai Huang, Sega Huang, Ping Kuan Hsieh, Shang Ru Wu, Yue Yun Chang, Jiming Liu, Po Shin Peng, Chia Yi Lin. Novel recombinant rabbit monoclonal antibodies for cancer biology research [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1325.