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

Background/Aim: A multistep sorting method for enrichment of rare cells, such as circulating tumor cells, in the blood without cumbersome pretreatments required by most flow cytometry-based methods, which lead to high cost and decreased detection efficiency, was developed. Materials and Methods: After only hemolysis and cell staining, cancer cells are enriched by repetitive sorting (3x) based on nuclear-positive, cytokeratin-positive, and CD45-negative expression. Results: Experiments using spikes of PC-9 cells showed a mean recovery of 65% and mean purity of 83%, which was retained up to 72 hours after blood draw using preservative tubes. Significant differences in expression level of programmed death-ligand 1 or vimentin were observed between high-and low-expressing cell lines, concurrently with enrichment. Next-generation sequencing analysis of recovered PC-9, A549, and MDA-MB231 cells successfully detected all known mutations. Conclusion: This novel isolation method applicable for preserved samples with sufficient recovery and purity may be substantially beneficial for recovering cells for subsequent molecular analysis.