Abstract
Conformational antibodies specific for amyloid-forming peptides and proteins are important for a range of biomedical applications, including detecting, inhibiting, and potentially treating protein aggregation disorders ranging from Alzheimer’s to Parkinson’s diseases. Generation of anti-amyloid antibodies is greatly complicated by the complex, heterogeneous and insoluble nature of amyloid antigens. Here we describe systematic methods for isolating and affinity maturing anti-amyloid antibodies using yeast surface display. Magnetic-activated cell sorting is used to sort single-chain antibody libraries positively for binding to amyloid antigens and negatively against the corresponding disaggregated antigens to remove antibodies that bind in a conformation-independent manner. Isolated lead antibody clones with conformational specificity are affinity matured via targeted CDR mutagenesis and magnetic-activated cell sorting.
Alec A. Desai and Jennifer M. Zupancic are co-first authors.
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Acknowledgments
We thank the Tessier lab for their feedback on the manuscript. This work was supported by the National Institutes of Health (RF1AG059723, R01AG050598, and R35GM136300 to P.M.T.) and National Science Foundation (CBET 1159943, 1605266, and 1813963 to P.M.T., Graduate Research Fellowship to M.D.S.), the Albert M. Mattocks Chair (to P.M.T).
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Desai, A.A., Zupancic, J.M., Smith, M.D., Tessier, P.M. (2022). Isolating Anti-amyloid Antibodies from Yeast-Displayed Libraries. In: Traxlmayr, M.W. (eds) Yeast Surface Display. Methods in Molecular Biology, vol 2491. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2285-8_22
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DOI: https://doi.org/10.1007/978-1-0716-2285-8_22
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