Ultrahigh-Throughput Enrichment of Circulating Tumor Cells from Whole Blood

Circulating tumor cells (CTCs) in peripheral blood serve as valuable indicators of cancer progression and important biomarkers for the early diagnosis and prognosis of the disease. Unfortunately, CTCs are typically present at extremely low concentrations, and the challenge of enriching and isolating rare cells is often insurmountable. Herein, we present a fluorescence-based active cell sorting system designed to isolate rare cells from peripheral blood. Our platform is based on the principle of positive selection, where cell-surface markers are specifically labelled with fluorescent antibodies, ranked by aliquots, and subsequently sorted. The developed system comprises a multilayer microfluidic device that incorporates 10 parallel channels equipped with pneumatically actuated control valves. A one-dimensional complementary metal-oxide-semiconductor image sensor integrated with a field-programmable gate array enables real-time fluorescence detection across all channels, with the system processing 0.5 mL of minimally processed whole blood per minute. The platform is validated by spiking cancer cells into 5-fold diluted, minimally processed whole blood, at a ratio of 1:10⁸ cancer cells to blood cells. Remarkably, after only three rounds of sorting, cancer cells are enriched by 8 orders of magnitude with a recovery rate exceeding 90%. To showcase the utility of the platform as a clinical tool, we assay clinical blood samples from cancer patients, successfully isolating CTCs at an average concentration of 7.6 CTCs/mL. Such performance metrics confirm immediate utility in the enrichment of CTCs and as a minimally invasive biopsy tool in clinical diagnostics and cancer research.