Abstract 2588: Bioengineering of site-specific antibody labeling from basic research to spatial omics platforms

Abstract Over the last few years, spatial multi-omics platforms have emerged as a powerful tool for characterizing cancer biology in multiple dimensions. Notably, these systems typically rely on multiplex immunolabeling. Conventionally, multiplexing is accomplished through carefully selecting primary antibodies from different animal species, and in combination with their corresponding polyclonal secondary antibodies. However, given the practical complication related to the species availability of primary antibodies, the multiplexing labeling is still under constraint due to the necessity for matched secondary species labels and the availability. For decades, non-specific labeling of primary antibodies through amine- or thiol-reactive chemistries was widely performed. Evolving research and commercial products have sought to overcome this antibody-based multiplexing limitations by developing enzymatic reaction-directed chemical reaction or antibody targeting proteins that could contain affinity to the Fc or Fab regions of antibodies. However, these methods mentioned above have substantive limitations and side-effects on affinity, conjugation number, buffer incompatibilities, storage incompatibilities, insufficient conjugation, and uncontrolled stoichiometric labeling. Herein, we introduce an efficient, sustainable, reproducible site-specific antibody covalent labeling strategy through engineering anti-IgG secondary nanobodies. These infer the stability, high-affinity of species specific and easy-to-produce IgG binders with the permanence of controlled activation for proximity-driven intermolecular covalent conjugation, with a promise for efficient multiplexity. This labeling technology addresses antibody labeling challenges through currently available approaches, and would substantially enhance multiplexity in a wide variety of research and clinical settings. Citation Format: Kai Tao, Matthew J. Rames, MingChong Dai, Gulsu Sener, Samuel Olson, Srivathsan Ranganathan, Sila Ozdemir, Adam Oken, Michael Brasino, Randy Armstrong, Yu-Jui Chiu, Ece Eksi, Bruce Branchaud, Sadik Esener. Bioengineering of site-specific antibody labeling from basic research to spatial omics platforms [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2588.