Cellular barcodes for efficiently profiling single-cell secretory responses by microengraving.

We present a method that uses fluorescent cellular barcodes to increase the number of unique samples that can be analyzed simultaneously by microengraving, a nanowell array-based technique for quantifying the secretory responses of thousands of single cells in parallel. Using n different fluorescent dyes to generate 2(n) unique cellular barcodes, we achieved a 2(n)-fold reduction in the number of arrays and quantity of reagents required per sample. The utility of this approach was demonstrated in three applications of interest in clinical and experimental immunology. Using barcoded human peripheral blood mononuclear cells and T cells, we constructed dose-response curves, profiled the secretory behavior of cells treated with mechanistically distinct stimuli, and tracked the secretory behaviors of different lineages of CD4(+) T helper cells. In addition to increasing the number of samples analyzed by generating secretory profiles of single cells from multiple populations in a time- and reagent-efficient manner, we expect that cellular barcoding in combination with microengraving will facilitate unique experimental opportunities for quantitatively analyzing interactions among heterogeneous cells isolated in small groups (~2-5 cells).