Expediting Antibody Discovery and Characterization with Flow Cytometry

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Recent advances in analytical techniques have increased the efficiency of therapeutic antibody discovery and facilitated rapid growth of the mono-clonal antibody (mAb) market over the past 25 years. There are now over 100 FDA approved mAb therapeutics, with the vast majority indicated for treatment of cancers, for example Rituximab treatment of non-Hodgkin lymphoma, and immune disorders, such as Humira treatment of rheumatoid arthritis1.

Next generation mAbs utilize the backbone of existing therapeutics with modifications to improve characteristics, such as potency or specificity. These can be categorized into four main groups based on the modifications they include2. The first group have engineered Fc regions to enhance function and/or half-life. The second have adaptations to improve specificity, including bi- and tri-specific antibodies. The third group are fragmented antibodies such as nanobodies, with greater stability and tissue penetration compared to native immunoglobulins3. The final category are conjugated antibodies, which includes antibody-drug conjugates.

Conventional methods for characterizing antibody binding and function, such as ELISA or traditional flow cytometry, can suffer from limited throughput, large sample volume requirements and the need to combine results from multiple instruments. The iQue® Advanced Flow Cytometry Platform, with validated assays and reagent kits, provides a streamlined solution for high-throughput antibody screening assays. Low volume samples are rapidly acquired from 96- or 384-well plates, then processed in the integrated iQue Forecyt® software, with auto-compensation and simple data analysis to deliver fast, actionable results.

This eBook highlights some of the most recent advancements in the field of antibody discovery, covering a range of methods of antibody characterization and the novel modalities produced. One article outlines work from scientists at The University of California, who used artificial intelligence to predict binding affinity from antibody sequence. Another article describes a study from the University of Southampton, which suggests fine-tuning antibody binding affinity may result in improved efficacy, with some lower affinity immunomodulatory antibodies mediating greater signalling. The included application notes from Sartorius detail in vitro assays that use the iQue® platform for functionally profiling antibodies. This collection of articles outlines the use of flow cytometry as a critical tool to advance antibody discovery.



  1. H. Kaplon, A. Chenoweth, S. Crescioli & J. M. Reichert, Antibodies to watch in 2022, mAbs, 14:1 2022.
  2. C. Challener, Witnessing Major Growth in Next-Generation Antibodies, BioPharm International, 30 (4) 2017.
  3. S. Sun et al., Nanobody: A Small Antibody with Big Implications for Tumor Therapeutic Strategy, Int J Nanomedicine, 22;16:2337-2356. 2021


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