Interfacing droplet microfluidics with antibody barcodes for multiplexed single-cell protein secretion profiling.

Multiplexed protein secretion analysis of single cells is important to understand the heterogeneity of cellular functions and processes in healthy and disease states. However, current single-cell platforms, such as microwell-, microchamber-, or droplet-based assays, suffer from low single-cell occupancy, waste of reagents, limited sensitivity, or inability to perform necessary operations, etc. To overcome these drawbacks, we present an integrated droplet microfluidic device that interfaces with spatially patterned antibody barcodes for multiplexed single-cell secretome analysis. The trapping array of 100 picoliter-sized isolation chambers could achieve >80% single-cell capture efficiency with >90% viability. The single-cell analysis microchip was validated by the detection of four-plexed cytokines, including IL-8, MCP-1, MIP-1b, and TNF-a/IL-10, from unstimulated and lipopolysaccharide (LPS)-stimulated individual human macrophages. We also successfully applied the platform to profile protein secretions of human tumor cell lines and primary/metastatic cancer cells dissociated from cancer patients to observe the secretion heterogeneity among cells. This unique microfluidic platform enables multiplexed secretion assays for static droplet microfluidics, provides a reliable and straightforward workflow for protein secretion assays based on a low number of single cells in a short incubation time (∼4 h), and could have widespread applications for studying secretion-mediated cellular heterogeneity.

Enrichment and single-cell analysis of circulating tumor cells.

Up to 90% of cancer-related deaths are caused by metastatic cancer. Circulating tumor cells (CTCs), a type of cancer cell that spreads through the blood after detaching from a solid tumor, are essential for the establishment of distant metastasis for a given cancer. As a new type of liquid biopsy, analysis of CTCs offers the possibility to avoid invasive tissue biopsy procedures with practical implications for diagnostics. The fundamental challenges of analyzing and profiling CTCs are the extremely low abundances of CTCs in the blood and the intrinsic heterogeneity of CTCs. Various technologies have been proposed for the enrichment and single-cell analysis of CTCs. This review aims to provide in-depth insights into CTC analysis, including various techniques for isolation of CTCs with capture methods based on physical and biochemical principles, and single-cell analysis of CTCs at the genomic, proteomic and phenotypic level, as well as current developmental trends and promising research directions.

Determination of cysteine and glutathione based on the inhibition of the dinuclear Cu(II)-catalyzed luminol-H2O2 chemiluminescence reaction.

The catalyzed luminol chemiluminescent reaction has received a great amount of attention because of its high sensitivity and low background signal which make the reaction an attractive analytical chemistry tool. The present study, introduces the beneficial catalytic effects of dinuclear Cu(II) complex [Cu2L2(TAE)]X2, where TAE=tetraacetylethane; L=N,N(')-dibenzylethylenediamine and X=ClO4 on the luminol chemiluminescent reaction as a novel probe for the determination of glutathione (GSH) and L-cysteine (CySH) in human serum and urine. The [Cu2L2(TAE)]X2 has exhibited highly efficient catalytic activity of luminol CL as an artificial peroxidase model at pH as low as 7.5 in water in the presence of H2O2⋅GSH and CySH can induce a sharp decrease in CL intensity from the [Cu2L2(TAE)]X2-catalyzed luminol system. Under the selected experimental conditions, a linear relationship was obtained between the CL intensity and the concentrations of GSH and CySH in the range of 1.0×10(-7)-1.0×10(-4) M, with detection limits (S/N=3) of 2.7×10(-8) and 6.8×10(-8) M and RSD<4.2% (n=7) for GSH and CySH, respectively.