Cell-tracking Raman-activated cell sorting

Summary Raman-activated cell sorting (RACS) integrates Raman spectroscopy with cell sorting to isolate cells based on intrinsic molecular composition in a label-free manner. However, its adoption has been limited because long-term operational stability is difficult to maintain. The intrinsically weak Raman scattering cross section requires long acquisition times (∼100 ms) per cell, constraining extended operation. Earlier attempts to address this have relied on trap-and-release strategies, which demand complex fluidic control and compromise robustness. We present a trap-free, continuous-flow, cell-tracking RACS platform in which the excitation beam is synchronously scanned with individual cells as they traverse a microfluidic channel, enabling stable Raman spectral acquisition during prolonged operation. This system achieved sustained sorting of triacylglycerol-rich Saccharomyces cerevisiae cells for over 4 h with ∼90% purity, screening over 30,000 cells. These results establish a robust, scalable RACS architecture that enables large-scale, label-free functional screening, directed evolution, and metabolic engineering.