Toward Rare Blood Cell Preservation for RNA Sequencing

Area of Science:

• Molecular Biology
• Genomics
• Biotechnology

Background:

• Cancer progression involves complex cellular events requiring molecular analysis.
• Next-generation sequencing necessitates high-quality nucleic acids from human samples.
• Existing RNA preservation methods have limitations for low blood volumes and long-term storage.

Purpose of the Study:

• To develop a robust protocol for blood preservation and long-term storage that maintains RNA integrity.
• To assess the protocol's suitability for preserving rare cell samples, such as circulating tumor cells.
• To ensure gene expression and cell surface antigen availability after preservation.

Main Methods:

• Development of a novel blood preservation protocol.
• Evaluation of RNA integrity using RNA integrity number (RIN) values.
• Assessment of gene expression stability using correlation coefficients (r²).
• Analysis of cell surface antigen availability for antibody selection.
• Data mining for fusion events to detect rare tumor cells.

Main Results:

• The developed protocol maintained RNA integrity (RIN > 8) and gene expression stability (r² > 0.88) after preservation and long-term storage (r² = 0.95).
• Cell surface antigens remained available for antibody selection (r² = 0.95).
• Rare tumor cells were detectable among background blood cells using data mining for fusion events, irrespective of fixation.

Conclusions:

• The developed protocol is effective for blood preservation and long-term storage, ensuring high-quality RNA for downstream applications like RNA sequencing.
• This method is suitable for preserving rare blood cells, including circulating tumor cells, enabling their analysis.
• The protocol supports comprehensive molecular profiling of blood samples, advancing cancer research and diagnostics.