Simplified Drop-seq workflow with minimized bead loss using a bead capture and processing microfluidic chip

Single-cell RNA-sequencing (scRNA-seq) has revolutionized biomedical research by enabling the in-depth analysis of cell-to-cell heterogeneity of tissues with unprecedented resolution. One of the catalyzing technologies is single cell droplet microfluidics, which has massively increased the overall cell throughput, routinely allowing the analysis of thousands of cells per experiment at a relatively low cost. Among several existing droplet-based approaches, the Drop-seq platform has emerged as one of the most widely used systems. Yet, this has surprisingly not incentivized major refinements of the method, thus restricting any lab implementation to the original Drop-seq setup, which is known to suffer from up to 80% bead loss during the process.

Researchers from Ecole Polytechnique Fédérale de Lausanne (EPFL) present a systematic re-engineering and optimization of Drop-seq: first, they re-designed the original dropleting device to be compatible with both air-pressure systems and syringe pumps, thus increasing the overall flexibility of the platform. Second, they devised an accompanying chip for post-encapsulation bead processing, which simplifies and massively increases Drop-seq’s cell processing efficiency. Taken together, the presented optimization efforts result in a more flexible and efficient Drop-seq version.

Overview of the revised Drop-seq workflow

The revised work-flow (dashed lines) is a flexible protocol that simplifies handling and increases the efficiency of the original protocol (solid lines). a) By redesigning the original chip into the e-chip, it is now possible to use either syringe pumps or an air-pressure regulator for the cell-bead encapsulation process. b) Utilizing a new bead capture and processing chip, the cp-chip, beads can now be either captured post droplet breakage, or directly captured from droplets for smaller bead quantities. c) Following the bead capture, it is possible to either retrieve the beads into a tube for the STAMP generation process (reverse transcription and exonuclease treatment), or to perform STAMP generation on-chip to further improve the overall bead recovery efficiency.