High-throughput single-cell activity-based screening and sequencing of antibodies using droplet microfluidics.,Nature Biotechnology

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High-throughput single-cell activity-based screening and sequencing of antibodies using droplet microfluidics.
Nature Biotechnology ( IF 33.1 ) Pub Date : 2020-03-30 , DOI: 10.1038/s41587-020-0466-7
Annabelle Gérard ₁ , Adam Woolfe ₁ , Guillaume Mottet ₂ , Marcel Reichen ₁ , Carlos Castrillon _{2,

3,

4} , Vera Menrath ₁ , Sami Ellouze ₁ , Adeline Poitou ₁ , Raphaël Doineau _{1,

3,

4,

5} , Luis Briseno-Roa _{1,

6} , Pablo Canales-Herrerias _{2,

3,

7} , Pascaline Mary ₈ , Gregory Rose ₈ , Charina Ortega ₈ , Matthieu Delincé ₈ , Sosthene Essono ₈ , Bin Jia _{9,

10} , Bruno Iannascoli ₂ , Odile Richard-Le Goff ₂ , Roshan Kumar ₈ , Samantha N Stewart ₈ , Yannick Pousse ₁ , Bingqing Shen ₁ , Kevin Grosselin _{1,

4} , Baptiste Saudemont _{2,

4} , Antoine Sautel-Caillé ₄ , Alexei Godina ₄ , Scott McNamara ₁ , Klaus Eyer _{7,

11} , Gaël A Millot ₁₂ , Jean Baudry ₇ , Patrick England _{13,

14} , Clément Nizak ₄ , Allan Jensen _{1,

15} , Andrew D Griffiths ₄ , Pierre Bruhns ₂ , Colin Brenan _{1,

8,

16}

Affiliation

HiFiBiO Therapeutics SAS, Paris, France.
Unit of Antibodies in Therapy and Pathology, Institut Pasteur, UMR1222 INSERM, Paris, France.
École Doctorale Frontières du Vivant (FdV), Centre de Recherches Interdisciplinaires, Paris, France.
Laboratoire de Biochimie (LBC), ESPCI Paris, PSL Research University, CNRS UMR8231 Chimie Biologie Innovation, Paris, France.
Institute for Integrative Nanosciences, Leibniz IFW Dresden e.V., Dresden, Germany.
Medetia Pharma, Institut Imagine, Paris, France.
Laboratoire Colloïdes et Matériaux Divisés (LCMD), ESPCI Paris, PSL Research University, CNRS UMR8231 Chimie Biologie Innovation, Paris, France.
HiFiBiO Therapeutics Inc., Cambridge, MA, USA.
Pfizer, Cambridge, MA, USA.
Abbvie Bioresearch Center, Worcester, MA, USA.
Laboratory for Functional Immune Repertoire Analysis, Institute of Pharmaceutical Sciences, D-CHAB, ETH Zürich, Zurich, Switzerland.
Institut Pasteur, Hub Bioinformatique et Biostatistique, C3BI, USR 3756 IP CNRS, Paris, France.
Plateforme de Biophysique Moléculaire, Institut Pasteur, CNRS UMR3528, Paris, France.
Centre d'Innovation et de Technologie, Plateforme de Biophysique des Macromolecules et de leurs Interactions, Institut Pasteur, Paris, France.
Lundbeck A/S, Valby, Denmark.
1CellBiO Inc., Watertown, MA, USA.

Mining the antibody repertoire of plasma cells and plasmablasts could enable the discovery of useful antibodies for therapeutic or research purposes1. We present a method for high-throughput, single-cell screening of IgG-secreting primary cells to characterize antibody binding to soluble and membrane-bound antigens. CelliGO is a droplet microfluidics system that combines high-throughput screening for IgG activity, using fluorescence-based in-droplet single-cell bioassays2, with sequencing of paired antibody V genes, using in-droplet single-cell barcoded reverse transcription. We analyzed IgG repertoire diversity, clonal expansion and somatic hypermutation in cells from mice immunized with a vaccine target, a multifunctional enzyme or a membrane-bound cancer target. Immunization with these antigens yielded 100-1,000 IgG sequences per mouse. We generated 77 recombinant antibodies from the identified sequences and found that 93% recognized the soluble antigen and 14% the membrane antigen. The platform also allowed recovery of ~450-900 IgG sequences from ~2,200 IgG-secreting activated human memory B cells, activated ex vivo, demonstrating its versatility.

中文翻译：

使用液滴微流控技术基于抗体的高通量单细胞活性筛选和测序。

挖掘浆细胞和浆母细胞的抗体库可以发现用于治疗或研究目的的有用抗体1。我们提出了一种高通量，分泌IgG的原代细胞单细胞筛选的方法，以表征抗体与可溶性和膜结合抗原的结合。CelliGO是一种液滴微流控系统，结合了高通量的IgG活性筛选（使用基于荧光的液滴内单细胞生物测定法2）和配对的抗体V基因测序（液滴内单细胞条形码反转录）。我们分析了用疫苗靶标，多功能酶或膜结合癌靶标免疫的小鼠的细胞中IgG的库多样性，克隆扩增和体细胞超突变。用这些抗原免疫产生每只小鼠100-1,000个IgG序列。我们从确定的序列中生成了77个重组抗体，发现93％的人识别了可溶性抗原，而14％的人识别了膜抗原。该平台还允许从体内分泌的约2,200 IgG分泌的活化人记忆B细胞中回收约450-900 IgG序列，这证明了其多功能性。

更新日期：2020-04-24

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