Fully Automated Continuous Centrifugal Microfluidics Isolates Natural Killer Cells with High Performance and Minimal Stress

Naturalkiller (NK) cells are a part of the innate immune system,providing the first line of defense against cancer cells and pathogensat an early stage. Hence, they are attracting attention as a valuableresource for allogeneic cell immunotherapy. However, NK cells existwith limited proportion in blood, and obtaining sufficient clinical-gradeNK cells with highly viable and minimal stress is critical for successfulimmune cell therapy. Conventional purification methods via immunoaffinityor density gradient centrifugation had several limitations in yield,purity, and cellular stress, which might cause an increased risk forgraft versus host disease and reduced efficacy due to NK cell malfunction,exhaustion, and apoptosis. Moreover, reducing the variations of isolationperformance caused by the manual process is another unmet need foruniform quality of the living drug. Here, an automated system usingan NK disc (NKD) based on continuous centrifugal microfluidics (CCM)technology was developed to isolate NK cells from whole blood withhigh yield, purity, reproducibility, and low stress. The CCM technology,which operates fluidic manipulation under disc rotation, enabled preciseextraction of the ultra-thin target fluid layer generated by bloodcentrifugation. Compared to the conventional manual method, the CCM-NKDisolated NK cells with higher yield (recovery rate) and purity, whilemaintaining better reproducibility. Furthermore, since the CCM-NKDmaintained substantially milder centrifugation conditions (120 xg for 10 min) compared to the conventional approach (1200xg for 20 min), it showed reduced cellular stressand increased antioxidant capacity in the isolated NK cells. Basedon the results, the CCM-NKD is expected to be a useful tool to providehighly intact and viable cell weapons for successful immune cell therapy.