Isolating and cryopreserving pig skin cells for single-cell RNA sequencing study

Area of Science:

• Dermatology
• Genomics
• Biotechnology

Background:

• Pig skin shares architectural and physiological similarities with human skin, making it a valuable model for research.
• Single-cell RNA sequencing (scRNA-seq) offers high-resolution insights into cellular composition and function.
• Applying scRNA-seq to pig skin research is currently limited due to challenges in obtaining quality single cells.

Purpose of the Study:

• To establish a reliable method for isolating and cryopreserving high-quality single cells from pig skin for scRNA-seq.
• To validate the efficacy of the developed protocol in maintaining cell viability and gene expression profiles.

Main Methods:

• Utilized the Miltenyi Human Whole Skin Dissociation kit and gentleMACS Dissociator for efficient tissue dissociation.
• Optimized a cryopreservation technique using 90% FBS + 10% DMSO for long-term storage of single cells.
• Analyzed cell viability, aggregation, and gene expression profiles post-cryopreservation.

Main Results:

• Successfully dissociated pig skin into single cells with high viability.
• Demonstrated that cryopreservation using FBS and DMSO did not negatively impact cell integrity or gene expression.
• Confirmed the protocol's ability to yield high-quality single cells suitable for identifying major skin cell types via scRNA-seq.

Conclusions:

• The developed protocol provides a robust and reproducible method for preparing pig skin single cells for scRNA-seq.
• This advancement facilitates deeper investigation into pig skin biology and therapeutic testing using scRNA-seq.
• The protocol is a valuable resource for the broader skin research community.