Issue 8, 2022

IL-2 secretion-based sorting of single T cells using high-throughput microfluidic on-cell cytokine capture

Abstract

Secreted proteins are critical for the coordination of potent immune defenses, such as in engineered T cell therapies, however, there are few widely accessible approaches to accurately analyze and sort large numbers of cells based on their secretory functions. We report a workflow for the rapid screening and sorting of single individual T cells based on IL-2 secretion accumulated at high concentrations in nanoliter droplets and encoded back onto the secreting cell's surface. In our method, droplets are used solely to partition cells, enabling rapid accumulation of signals onto cell surfaces, and eliminating diffusive crosstalk between neighbors. All downstream sorting leverages conventional high-throughput and readily accessible flow cytometry after the emulsion is disrupted. We achieve monodisperse droplet generation (CV < 10%) at flow rates up to 200 μL min−1 using step emulsification, enabling processing of entire libraries of cells within tens of minutes without significant secretion crosstalk. In comparison to our approach, strong mitogenic activation overwhelmed the conventional bulk on-cell cytokine assay, rendering labeled, non-activated cells indistinguishable from actively secreting neighbors within one hour. Processing of identical cell mixtures following droplet encapsulation yielded no apparent crosstalk even after three hours. Instead, IL-2 production spanning several orders of magnitude was observed from roughly 20% of analyzed activated lymphocytes, representing an at least 10-fold increase in dynamic range compared to unencapsulated cells. Secreting cells could also be sorted using fluorescence activated cell sorting (FACS). The approach can ultimately enable sorting of cells based on functional properties with higher accuracy in a more accessible format to life science researchers.

Graphical abstract: IL-2 secretion-based sorting of single T cells using high-throughput microfluidic on-cell cytokine capture

Supplementary files

Article information

Article type
Paper
Submitted
04 دسمبر 2021
Accepted
08 مارٕچ 2022
First published
09 مارٕچ 2022

Lab Chip, 2022,22, 1576-1583

IL-2 secretion-based sorting of single T cells using high-throughput microfluidic on-cell cytokine capture

R. Dimatteo and D. Di Carlo, Lab Chip, 2022, 22, 1576 DOI: 10.1039/D1LC01098K

To request permission to reproduce material from this article, please go to the Copyright Clearance Center request page.

If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given.

If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page.

Read more about how to correctly acknowledge RSC content.

Social activity

Spotlight

Advertisements