Antibody specificity is arguably the most important and most challenging property to optimize during <i>in vitro</i> antibody selection and affinity maturation. We have sought to develop methods for both designing and sorting antibody libraries that maximize the specificity of the selected variants. First, we have developed a novel library design method (Natural Diversity Mutagenesis) that samples wild-type residues and a small number of the most frequently occurring residues at each position of antibody complementarity-determining regions (CDRs) and/or framework regions based on natural antibody diversity. This mutagenesis approach results in antibody libraries with variants that have a wide range of numbers of mutations, including antibodies with single mutations and others with tens of mutations. Second, we have developed methods for performing strong negative selections using different types of non-specific (peptide and protein) reagents and strong positive selections using the targets of interest. Interestingly, we have identified reagents that are unusually effective at removing non-specific antibody variants during negative selections. Moreover, we find that our library design and selection methods result in the identification of antibody variants with extremely high specificity in addition to high expressibility and solubility. These variants are enriched with specific types of amino acids in their CDRs that are linked to drug-like antibody properties. We will discuss how we are using these and related approaches to improve <i>in vitro</i> isolation of antibodies with properties that rival those of natural antibodies.

Improved library design and selection methods for identifying antibodies with high specificity and solubility

255th National Meeting (2018)

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