Cell motility influences microfluidics capturing in scRNA-seq.

Area of Science:

• Molecular Biology
• Genomics
• Biotechnology

Background:

• Microfluidic platforms for single-cell RNA sequencing (scRNA-seq) are typically designed for immotile cells.
• Motile cells, possessing flagella or cilia, present unique challenges for microfluidic isolation and encapsulation.
• Existing scRNA-seq protocols may introduce biases due to the loss of motile subpopulations.

Purpose of the Study:

• To investigate the encapsulation efficiency of motile cells, specifically Trypanosoma brucei, on the 10x Genomics platform.
• To develop and validate an optimized protocol for scRNA-seq of motile cells, ensuring representative data capture.
• To address the limitations of current microfluidic systems in handling non-standard mammalian cells.

Main Methods:

• Studied encapsulation efficiency of Trypanosoma brucei at room temperature on the 10x Genomics platform.
• Implemented a rapid cooling protocol to 0°C prior to cell encapsulation.
• Analyzed scRNA-seq data to assess representativeness and identify biases.

Main Results:

• Maintaining Trypanosoma brucei at room temperature resulted in low encapsulation yield.
• Rapid cooling to 0°C significantly reduced parasite motility and prevented transcriptomic alterations.
• The optimized protocol enabled the generation of a representative scRNA-seq dataset, avoiding loss of motile forms.

Conclusions:

• Optimized protocols are crucial for obtaining representative scRNA-seq data from motile cells.
• Rapid cooling is an effective strategy to mitigate motility-associated biases in microfluidic cell encapsulation.
• This work underscores the need for tailored approaches for non-standard mammalian cells in scRNA-seq applications.