High-purity Isolation for Genotyping Rare Cancer Cells from Blood Using a Microfluidic Chip Cell Sorter.

Area of Science:

• Biotechnology
• Cell Biology
• Oncology

Background:

• Traditional flow cytometry methods for rare cell isolation are costly and inefficient.
• Cumbersome pretreatments reduce detection efficiency for circulating tumor cells (CTCs).

Purpose of the Study:

• To develop a simplified, cost-effective multistep sorting method for enriching rare cells, like CTCs, from blood.
• To improve the efficiency and accessibility of rare cell isolation for subsequent molecular analysis.

Main Methods:

• A multistep sorting technique involving hemolysis and cell staining.
• Repetitive sorting (3×) based on nuclear-positive, cytokeratin-positive, and CD45-negative expression for cancer cell enrichment.
• Utilized preservative tubes for blood samples, maintaining integrity up to 72 hours post-collection.

Main Results:

• Achieved a mean recovery of 65% and mean purity of 83% for spiked PC-9 cells.
• Detected significant differences in programmed death-ligand 1 and vimentin expression between cell lines post-enrichment.
• Successfully identified all known mutations in recovered cancer cell lines (PC-9, A549, MDA-MB231) via next-generation sequencing.

Conclusions:

• The novel isolation method offers sufficient recovery and purity for preserved samples.
• This technique is beneficial for recovering rare cells for downstream molecular profiling.
• Simplifies rare cell isolation, potentially enhancing cancer diagnostics and research.