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On-Chip Analysis of Protein Secretion from Single Cells Using Microbead Biosensors
Diana F Cedillo-Alcantar1, Roberto Rodriguez-Moncayo1, Jose L Maravillas-Montero2
1Laboratory of Microtechnologies for Biomedicine, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional (Cinvestav), Monterrey 66628, Nuevo León Mexico.
Insights
This study introduces a microfluidic device for quantifying cytokine secretion from single immune cells. The platform enables precise measurement of interleukin-8 (IL-8) from monocytes, revealing insights into cellular function heterogeneity.
Area of Science:
- Immunology
- Biotechnology
- Microfluidics
Background:
- Understanding single immune cell effector functions, like cytokine secretion, is crucial for immune system insights, diagnostics, and therapeutics.
- Current methods may lack the precision to analyze cytokine secretion at the single-cell level.
- Heterogeneity in cellular function is vital for applications in cancer research and drug discovery.
Purpose of the Study:
- To develop and validate a microfluidic device for high-throughput, single-cell cytokine secretion analysis.
- To quantitate cytokine secretion from individual immune cells, specifically monocytes.
- To investigate the impact of disrupted paracrine signaling on cytokine secretion.
Main Methods:
- A microfluidic device with 1008 microchambers (∼500 pL each) was designed to pair single cells with antibody-functionalized microbeads.
- Hydrodynamic traps and integrated microvalves were used to isolate cell/bead pairs and prevent cross-contamination.
- A fluorescence sandwich immunoassay was implemented on microbeads for cytokine detection, achieving a 9 pg/mL limit of detection.
Main Results:
- The device successfully detected interleukin-8 (IL-8) secreted by single human monocytes in response to lipopolysaccharide (LPS).
- A limit of detection of 9 pg/mL for IL-8 was achieved using the fluorescence immunoassay.
- A significant reduction in IL-8-secreting monocytes was observed when paracrine signaling was disrupted.
Conclusions:
- The developed microfluidic platform enables precise quantification of single-cell cytokine secretion.
- This technology offers a valuable tool for studying cellular function heterogeneity in various biological contexts.
- Potential applications include cancer research, antibody discovery, and rare cell screening.
Abstract:
The profiling of the effector functions of single immune cells─including cytokine secretion─can lead to a deeper understanding of how the immune system operates and to potential diagnostics and therapeutical applications. Here, we report a microfluidic device that pairs single cells and antibody-functionalized microbeads in hydrodynamic traps to quantitate cytokine secretion. The device contains 1008 microchambers, each with a volume of ∼500 pL, divided into six different sections individually addressed to deliver an equal number of chemical stimuli. Integrating microvalves allowed us to isolate cell/bead pairs, preventing cross-contamination with factors secreted by adjacent cells. We implemented a fluorescence sandwich immunoassay on the biosensing microbeads with a limit of detection of 9 pg/mL and were able to detect interleukin-8 (IL-8) secreted by single blood-derived human monocytes in response to different concentrations of LPS. Finally, our platform allowed us to observe a significant decrease in the number of IL-8-secreting monocytes when paracrine signaling becomes disrupted. Overall, our platform could have a variety of applications for which the analysis of cellular function heterogeneity is necessary, such as cancer research, antibody discovery, or rare cell screening.
