Multiplexed functional macrophage profiling at single-cell resolution using Nanovial technology 2546

Description A pervading constituent of the tumor microenvironment, macrophages represent an attractive target for antitumor therapies and a candidate for engineered cell therapy. Therapeutic progress has been hindered by macrophage diversity underscoring the need for single-cell resolution approaches. To address this gap, we have developed a workflow utilizing Nanovials that integrates functional cytokine and protein phenotypes with gene expression patterns of individual cells in a single assay. Using M1 and M2 polarized human macrophages we compare intracellular cytokine staining with secreted protein signal. Unlike intracellularly stained cells, cells used in the Nanovial assay remain viable and were single-cell sequenced, pairing secreted protein with gene expression data. Leveraging a novel multicell Nanovial assay, we co-load activated macrophages with CD4 T Cells and assay cytokine secretion resulting from the interaction between activated macrophages and T cells. By utilizing barcoded Nanovials and cell hashtags we input co-loaded macrophage and T-cells into the single-cell sequencing workflow and analyzed the gene expression from both the macrophage and the T cell it was co-loaded with. This unprecedented insight into macrophage T-cell interactions can be utilized to facilitate researchers’ analysis of macrophages without the need for specialized equipment, precipitating a major advancement in the understanding of macrophage-based therapies.