Bumol, T. F. | Search bioRxiv

bioRxiv2020

Multi-omic Single-cell Atlas Reveals Broad Ex Vivo Effects on Human Immunobiology

Multi-omic profiling of human peripheral blood is increasingly utilized to identify biomarkers and pathophysiologic mechanisms of disease. The importance of these platforms in clinical and translational studies led us to investigate the impact of delayed blood processing on the numbers and state of peripheral blood mononuclear cells (PBMC) and on the plasma proteome. Similar to previous studies, we show minimal effects of delayed processing on the numbers and general phenotype of PBMCs up to 18 hours. In contrast, profound changes in the single-cell transcriptome and composition of the plasma proteome become evident as early as 6 hours after blood draw. These reflect patterns of cellular activation across diverse cell types that lead to progressive distancing of the gene expression state and plasma proteome from native in vivo biology. Differences accumulating during an overnight rest (18 hours) could confound relevant biologic variance related to many underlying disease states.

immunology↗

bioRxiv2020

Integrated single cell analysis of chromatin accessibility and cell surface markers

Single-cell measurements of cellular characteristics have been instrumental in understanding the heterogeneous pathways that drive differentiation, cellular responses to extracellular signals, and human disease states. scATAC-seq has been particularly challenging due to the large size of the human genome and processing artefacts resulting from DNA damage that are an inherent source of background signal. Downstream analysis and integration of scATAC-seq with other single-cell assays is complicated by the lack of clear phenotypic information linking chromatin state and cell type. Using the heterogeneous mixture of cells in human peripheral blood as a test case, we developed a novel scATAC-seq workflow that increases the signal-to-noise ratio and allows simultaneous measurement of cell surface markers: Integrated Cellular Indexing of Chromatin Landscape and Epitopes (ICICLE-seq). We extended this approach using a droplet-based multiomics platform to develop a trimodal assay to simultaneously measure Transcriptomic state (scRNA-seq), cell surface Epitopes, and chromatin Accessibility (scATAC-seq) from thousands of single cells, which we term TEA-seq. Together, these multimodal single-cell assays provide a novel toolkit to identify type-specific gene regulation and expression grounded in phenotypically defined cell types.

immunology↗