μCB-seq – microfluidic cell barcoding and sequencing for high-resolution imaging and sequencing of single cells

Single-cell RNA sequencing (scRNA-seq) enables the investigation of complex biological processes in multicellular organisms with high resolution. However, many phenotypic features that are critical to understanding the functional role of cells in a heterogeneous tissue or organ are not directly encoded in the genome and therefore cannot be profiled with scRNA-seq. Quantitative optical microscopy has long been a powerful approach for characterizing diverse cellular phenotypes including cell morphology, protein localization, and chemical composition. Combining scRNA-seq with optical imaging has the potential to provide comprehensive single-cell analysis, allowing for functional integration of gene expression profiling and cell-state characterization. However, it is difficult to track single cells through both measurements; therefore, coupling current scRNA-seq protocols with optical measurements remains a challenge.

Researchers from the University of California, Berkeley report microfluidic cell barcoding and sequencing (μCB-seq), a microfluidic platform that combines high-resolution imaging and sequencing of single cells. μCB-seq is enabled by a novel fabrication method that preloads primers with known barcode sequences inside addressable reaction chambers of a microfluidic device. In addition to enabling multi-modal single-cell analysis, μCB-seq improves gene detection sensitivity, providing a scalable and accurate method for information-rich characterization of single cells.

μCB-seq device design and workflow

(A) schematic of the microfluidic device with control valves in blue and flow layer in red. Cells are loaded into the cell inlet and reagent is introduced through the reagent inlet. The device processes 10 cells in 10 individual reaction lanes, each ending in an output port. Reverse-transcribed cDNA is recovered from output ports for all cells, pooled in a single tube for off-chip library preparation using the mcSCRB-seq protocol, and sequenced using next-generation sequencing platforms. (B) Detailed diagram of the imaging module showing the imaging chamber. The two isolation valves can be actuated to actively capture a cell of interest in the imaging chamber. (C) Detailed diagram of one reaction lane showing the lysis and RT modules separated by valves. The textured reaction chamber in the lysis module is preloaded with barcoded RT primers.

Chen TN, Gupta A, Zalavadia MD, Streets A. (2020) μCB-seq: microfluidic cell barcoding and sequencing for high-resolution imaging and sequencing of single cells Lab Chip [published online ahead of print]. [article]

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