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bioRxiv · 10.1101/2021.11.22.469484

Simplified, Shear Induced Generation of Double Emulsions for Robust Compartmentalization during Single Genome Analysis

Cowell, T. W.·Dobria, A.·Han, H.-S.
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Abstract

Drop microfluidics has driven innovations for high throughput, low input analysis techniques such as single-cell RNA-seq. However, the instability of single emulsion (SE) drops occasionally causes significant merging during drop processing, limiting most applications to single-step reactions in drops. Here, we show that double emulsion (DE) drops address this critical limitation and completely prevent content mixing, which is essential for single entity analysis. DEs show excellent stability during thermal cycling. More importantly, DEs undergo rupture into the continuous phase instead of merging, preventing content mixing and eliminating unstable drops from the downstream analysis. Due to the lack of drop merging, the monodispersity of drops is maintained throughout a workflow, enabling the deterministic manipulation of drops downstream. We also developed a simple, one-layer fabrication method for DE drop makers. This design is powerful as it allows robust production of single-core DEs at a wide range of flow rates and better control over the shell thickness, both of which have been significant limitations of conventional two-layer devices. This approach makes the fabrication of DE devices much more accessible, facilitating its broader adoption. Finally, we show that DE droplets effectively maintain the compartmentalization of single virus genomes during PCR-based amplification and barcoding, while SEs mixed contents due to merging. With their resistance to content mixing, DE drops have key advantages for multistep reactions in drops, which is limited in SEs due to merging and content mixing.

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Cowell, T. W., Dobria, A., Han, H.-S.. 2021-11-22. Simplified, Shear Induced Generation of Double Emulsions for Robust Compartmentalization during Single Genome Analysis. https://doi.org/10.1101/2021.11.22.469484

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