Functional single-cell hybridoma screening using droplet-based microfluidics

被引:201
作者
El Debs, Bachir [1 ,2 ,3 ]
Utharala, Ramesh [1 ]
Balyasnikova, Irina V. [4 ]
Griffiths, Andrew D. [2 ,3 ]
Merten, Christoph A. [1 ]
机构
[1] European Mol Biol Lab, Genome Biol Unit, D-69117 Heidelberg, Germany
[2] Univ Strasbourg, Inst Sci & Ingn Supramol, F-67083 Strasbourg, France
[3] Ctr Natl Rech Sci, Unite Mixte Rech 7006, F-67083 Strasbourg, France
[4] Univ Chicago, Brain Tumor Ctr, Chicago, IL 60637 USA
关键词
single-cell screening; high-throughput screening; cell-based assay; monoclonal antibody; angiotensin converting enzyme 1; ANGIOTENSIN-CONVERTING ENZYME; MONOCLONAL-ANTIBODIES; ENCAPSULATION; INHIBITORS;
D O I
10.1073/pnas.1204514109
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Monoclonal antibodies can specifically bind or even inhibit drug targets and have hence become the fastest growing class of human therapeutics. Although they can be screened for binding affinities at very high throughput using systems such as phage display, screening for functional properties (e. g., the inhibition of a drug target) is much more challenging. Typically these screens require the generation of immortalized hybridoma cells, as well as clonal expansion in microtiter plates over several weeks, and the number of clones that can be assayed is typically no more than a few thousand. We present here a microfluidic platform allowing the functional screening of up to 300,000 individual hybridoma cell clones within less than a day. This approach should also be applicable to nonimmortalized primary B-cells, as no cell proliferation is required: Individual cells are encapsulated into aqueous microdroplets and assayed directly for the release of antibodies inhibiting a drug target based on fluorescence. We used this system to perform a model screen for antibodies that inhibit angiotensin converting enzyme 1, a target for hypertension and congestive heart failure drugs. When cells expressing these antibodies were spiked into an unrelated hybridoma cell population in a ratio of 1:10,000 we observed a 9,400-fold enrichment after fluorescence activated droplet sorting. A wide variance in antibody expression levels at the single-cell level within a single hybridoma line was observed and high expressors could be successfully sorted and recultivated.
引用
收藏
页码:11570 / 11575
页数:6
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