The effects of cryopreservation on PBMCs transcriptome profile

Area of Science:

• Immunology
• Cell Biology
• Genomics

Background:

• Cryopreservation is crucial for storing biological specimens long-term.
• Optimizing cryopreservation minimizes damage and preserves cell function.
• Single-cell RNA sequencing (scRNA-seq) offers deeper insights into cellular heterogeneity.

Purpose of the Study:

• To optimize cryopreservation and recovery conditions for peripheral blood mononuclear cells (PBMCs).
• To evaluate the impact of cryopreservation on PBMC viability, population composition, and transcriptomic profiles using scRNA-seq.
• To assess the stability of PBMCs after 6 and 12 months of cryopreservation.

Main Methods:

• Peripheral blood mononuclear cells (PBMCs) were cryopreserved for up to 12 months.
• Cell viability was assessed using trypan blue and propidium iodide (PI) staining with FACS.
• Single-cell RNA sequencing (scRNA-seq) was performed on fresh and cryopreserved PBMCs to analyze cell populations and transcriptomes.

Main Results:

• Six major immune cell types were identified in both fresh and cryopreserved PBMCs.
• Cell viability remained stable for all immune cell types after 6 and 12 months of cryopreservation.
• A ~32% decline in scRNA-seq cell capture efficiency was observed after 12 months, with minimal transcriptomic changes (< two folds).

Conclusions:

• An optimized cryopreservation and recovery procedure effectively preserves PBMC viability and population composition for up to 12 months.
• While scRNA-seq cell capture efficiency decreases after prolonged cryopreservation, transcriptomic profiles remain largely unaffected.
• The optimized method ensures minimal impact on PBMC integrity and function for long-term storage.