Exceeding 80% Efficiency of Single-Bead Encapsulation in Microdroplets through Hydrogel Coating-Assisted Close-Packed...

Area of Science:

• Biotechnology
• Microfluidics
• Analytical Chemistry

Background:

• Droplet-based high-throughput analysis relies on efficient single microbead encapsulation.
• Poisson distribution of beads in droplets limits efficiency in assays like single-cell genomics.
• Existing methods like inertial ordering require advanced microfluidics and lack broad compatibility.

Purpose of the Study:

• To develop a general, simple, and compatible strategy to enhance microbead loading efficiency in microdroplets.
• To overcome the limitations imposed by random bead distribution and Poisson statistics.
• To improve the efficiency of droplet-based high-throughput assays.

Main Methods:

• Hydrogel coating-assisted close-packed ordering strategy for microbeads.
• Utilizing jetting microfluidics or vortex emulsification for hydrogel coating.
• Testing with 30-μm polystyrene beads and co-encapsulation with HEK293T cells and barcoded beads.

Main Results:

• Achieved over 80% bead-loading efficiency, experimentally determined as 81% for single 30-μm beads.
• Demonstrated strategy's robustness to bead polydispersity and raw bead type.
• Obtained a 68.8% cell capture rate in single-cell transcriptomics assays.
• Verified that reversible hydrogel coating does not impede RNA capture.

Conclusions:

• Hydrogel coating-assisted close-packed ordering is a simple and effective method to significantly improve microbead encapsulation efficiency.
• The strategy is compatible with diverse bead types and microfluidic setups.
• This approach has broad applicability for enhancing various droplet-based high-throughput analyses.