Rapid synthesis and screening of natively paired antibodies against influenza hemagglutinin stem via oPool+ display

Ouyang, W. O.; Lv, H.; Liu, W.; Mou, Z.; Lei, R.; Pholcharee, T.; Wang, Y.; Dailey, K. E.; Gopal, A. B.; Choi, D.; Ardagh, M. R.; Talmage, L.; Rodriguez, L. A.; Dai, X.; Wu, N. C.

doi:10.1101/2024.08.30.610421

bioRxiv · 10.1101/2024.08.30.610421

Rapid synthesis and screening of natively paired antibodies against influenza hemagglutinin stem via oPool+ display

Ouyang, W. O.·Lv, H.·Liu, W.·Mou, Z.·Lei, R.·Pholcharee, T.·Wang, Y.·Dailey, K. E.·Gopal, A. B.·Choi, D.·Ardagh, M. R.·Talmage, L.·Rodriguez, L. A.·Dai, X.·Wu, N. C.

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Abstract

Antibody discovery is crucial for developing therapeutics and vaccines as well as understanding adaptive immunity. However, the lack of approaches to synthesize antibodies with defined sequences in a high-throughput manner represents a major bottleneck in antibody discovery. Here, we presented oPool+ display, a high-throughput cell-free platform that combined oligo pool synthesis and mRNA display to rapidly construct and characterize many natively paired antibodies in parallel. As a proof-of-concept, we applied oPool+ display to probe the binding specificity of >300 uncommon influenza hemagglutinin (HA) antibodies against 9 HA variants through 16 different screens. Over 5,000 binding tests were performed in 3-5 days with further scaling potential. Follow-up structural analysis of two HA stem antibodies revealed the previously unknown versatility of IGHD3-3 gene segment in recognizing the HA stem. Overall, this study established an experimental platform that not only accelerate antibody characterization, but also enable unbiased discovery of antibody molecular signatures.

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Ouyang, W. O., Lv, H., Liu, W., Mou, Z., Lei, R., Pholcharee, T., Wang, Y., Dailey, K. E., Gopal, A. B., Choi, D., Ardagh, M. R., Talmage, L., Rodriguez, L. A., Dai, X., Wu, N. C.. 2024-08-30. Rapid synthesis and screening of natively paired antibodies against influenza hemagglutinin stem via oPool+ display. https://doi.org/10.1101/2024.08.30.610421

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