Multiplexed node-pore sensing for high-throughput, label-free single-cell detection

We present a multiplexed four-channel Node-Pore Sensing (NPS) platform in which each channel is encoded with a unique Manchester-coded sequence selected by combinatorial optimization for minimum aperiodic cross-correlation. A structural classifier based on the node-pore structure of each Gold code sequence achieves 99.3% single-channel identification accuracy on 1480 labeled experimental transit events, without template alignment or velocity correction. A linear support vector machine successfully distinguishes single-channel events from multi-channel coincidences, enabling label-free, high-throughput single-cell detection across all four channels. This multiplexed architecture provides a route to measuring cell mechanical properties, such as stiffness and recovery time, across parallel channels, extending NPS mechanophenotyping of primary human mammary epithelial cells toward the throughput needed for clinical study.