Microfluidic Droplet-SERS Platform for Single-Cell Cytokine Analysis via a Cell Surface Bioconjugation Strategy

A microfluidic-based surface-enhanced Raman scattering (SERS) platform for analyzing cytokines secreted by single cells is reported based on the elaborate bioconjugation of the immuno-sandwich complex on the probed cell surface. This platform integrates the dual functions of microfluidic droplet separation of single cells and SERS measurement. Two immune nanoprobes (capture probe and SERS probe) are introduced into a microfluidic droplet along with a single cell. They were anchored to the cell membrane protein surface by capturing secreted cytokines to form an immune sandwich structure, realizing the enrichment effect of cytokines above the cell membrane surface and the amplification effect of SERS detection probes. This single-cell analytical platform was applied to track specific cell-secreted vascular endothelial growth factor (VEGF) of different cell lines (MCF-7, SGC, and T24), and highly sensitive detection of VEGF was achieved. Chemometric methods (principal component analysis and t-distributed stochastic neighbor embedding) were adopted for the SERS data analysis, and the support vector machine (SVM) discriminant model was established to test the data. These chemometric methods successfully identify significant differences in the secreting ability of cytokines among three kinds of cancer cell lines, revealing cell heterogeneity. In addition, the behavior of single cells secreting VEGF was monitored time-dependently and was shown to increase with time. This work demonstrates the importance of tracking specific cells secreting cytokines based on the cell surface bioconjugation strategy. Our developed platform provides guidelines for using the single-cell exocytosis factors as biomarkers to assess the early diagnosis of cancer and provide physiological cues for learning single-cell secretions.