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Issue 45, 2020
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Rapid isolation of antigen-specific B-cells using droplet microfluidics

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Abstract

Monoclonal antibodies are powerful tools for scientific research and are the basis of numerous therapeutics. However, traditional approaches to generate monoclonal antibodies against a desired target, such as hybridoma-based techniques and display library methods, are laborious and suffer from fusion inefficiency and display bias, respectively. Here we present a platform, featuring droplet microfluidics and a bead-based binding assay, to rapidly identify and verify antigen-binding antibody sequences from primary cells. We used a defined mixture of hybridoma cells to characterize the system, sorting droplets at up to 100 Hz and isolating desired hybridoma cells, comprising 0.1% of the input, with a false positive rate of less than 1%. We then applied the system to once-frozen primary B-cells to isolate rare cells secreting target-binding antibody. We performed RT-PCR on individual sorted cells to recover the correctly paired heavy- and light-chain antibody sequences, and we used rapid cell-free protein synthesis to generate single-chain variable fragment-format (scFv) antibodies from fourteen of the sorted cells. Twelve of these showed antigen-specific binding by ELISA. Our platform facilitates screening animal B-cell repertoires within days at low cost, increasing both rate and range of discovering antigen-specific antibodies from living organisms. Further, these techniques can be adapted to isolate cells based on virtually any secreted product.

Graphical abstract: Rapid isolation of antigen-specific B-cells using droplet microfluidics

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Supplementary files

Article information


Submitted
15 May 2020
Accepted
30 Jun 2020
First published
20 Jul 2020

This article is Open Access

RSC Adv., 2020,10, 27006-27013
Article type
Paper

Rapid isolation of antigen-specific B-cells using droplet microfluidics

R. Ding, K. Hung, A. Mitra, L. W. Ung, D. Lightwood, R. Tu, D. Starkie, L. Cai, L. Mazutis, S. Chong, D. A. Weitz and J. A. Heyman, RSC Adv., 2020, 10, 27006
DOI: 10.1039/D0RA04328A

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