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Updated: Jun 20, 2026
Microfluidic Approach to Resolve Simultaneous and Sequential Cytokine Secretion of Individual Polyfunctional Cells
Published on: March 8, 2024
Detecting cytokine release from single T-cells
He Zhu1, Gulnaz Stybayeva, Jaime Silangcruz
1Biomedical Engineering, University of California, Davis, USA.
Insights
Researchers developed a microfluidic device to capture and analyze individual T-cells from human blood. This technology detects interferon-gamma production by single T-cells, offering new diagnostic potential for immune responses.
Area of Science:
- Biotechnology
- Immunology
- Microfluidics
Background:
- Cytokine production by leukocytes is crucial for immune response and has diagnostic value.
- Current methods for analyzing immune cells can be complex and require significant sample volumes.
Purpose of the Study:
- To develop a microfluidic device for capturing and analyzing single T-cells from minimally processed human blood.
- To detect interferon-gamma (IFN-gamma) production at the single-cell level.
Main Methods:
- Microfabricated surfaces with poly(ethylene glycol) (PEG) hydrogel microwells were created on glass slides.
- Antibodies for IFN-gamma and CD4 T-cells were immobilized within the microwells.
- A microfluidic device was used to isolate CD4 T-cells and detect IFN-gamma release after mitogenic activation.
Main Results:
- Individual CD4 T-cells were successfully isolated within PEG microwells from human blood.
- Interferon-gamma (IFN-gamma) cytokine signals were detected and colocalized with specific T-cells.
- The microfluidic system enabled analysis of single-cell immune responses.
Conclusions:
- The developed microfluidic device and process allow for high-density, single-cell analysis of immune cells.
- This technology holds promise for advancing diagnostic capabilities in immunology.
- Future work will focus on multiparametric functional analysis of immune cells.
Abstract:
The cytokine production by leukocytes correlates with body's ability to mount an immune response and therefore has high diagnostic value. In the present study we employed microfabricated surfaces to capture T-cells from minimally processed human blood, arrange these cells into a single cell array, and then detect interferon (IFN)-gamma released from individual cells. The fabrication of cell capture surfaces started with coating a silane-modified glass slide with a uniform layer of poly(ethylene glycol) (PEG) hydrogel. The hydrogel-coated slide was lyophilized and then incubated with a mixture of monoclonal anti-IFN-gamma and anti-CD4 antibodies (Abs). To define sites for single cell attachment, PEG hydrogel microwells (20 microm diameter) were photolithographically patterned on top of the Ab-containing hydrogel layer. This micropatterning process resulted in fabrication of PEG hydrogel microwells with Ab-decorated bottom and nonfouling walls. To minimize the blood volume requirement and to precisely define shear stress conditions, the engineered surface was enclosed inside a PDMS-based microfluidic device. Introduction of red blood cell (RBC) depleted whole human blood followed by controlled washing led to the isolation of individual CD4 T-cells within PEG microwells. Mitogenic activation and immunofluorescent staining performed inside the microfluidic chamber revealed IFN-gamma cytokine signal colocalized with specific T-cells. The device and process presented here will be expanded in the future to enable multiparametric functional analysis of immune cells organized into high density single cell arrays.
