Ultrasimple Single-Cell Detection of Multiple Cytokines by a Nanowell Chip Integrated with Encoded Microarrays

Area of Science:

• Immunology
• Biotechnology
• Materials Science

Background:

• Cytokine production signifies immune cell activation, but its analysis at the single-cell level is complex.
• Variations in cytokine secretion exist across and within immune cell phenotypes.
• Current methods require specialized laboratory equipment, limiting accessibility.

Purpose of the Study:

• To present a highly sensitive and robust method for quantifying cytokine expression at the single-cell resolution.
• To simplify multiplex single-cell cytokine analysis into an instrument-free, point-of-detection technology.

Main Methods:

• Development of a microchip featuring poly(dimethylsiloxane) nanowells on sticky tape.
• Integration of DNA-antibody convertible microarrays within each nanowell.
• Utilizing pipetting for cell loading, reagent addition, and sandwich enzyme-linked immunosorbent assay (ELISA) detection, with Fluorinert FC-40 for nanowell isolation.

Main Results:

• The assay demonstrated high sensitivity comparable to conventional methods, validated by measuring six recombinant cytokines.
• Successful validation through measuring cytokine production from hundreds of single cells.
• The method simplifies a complex multiplex single-cell assay.

Conclusions:

• The developed ultrasimple method enables instrument-free, single-cell cytokine quantification.
• This technology offers high sensitivity and robustness, potentially broadening its utility in clinical diagnostics.