Cellular Lensing and Near Infrared Fluorescent Nanosensor Arrays to Enable Chemical Efflux Cytometry

Abstract Nanosensor have proven to be powerful tools to monitor single biological cells and organisms, achieving spatial and temporal precision even at the single molecule level. However, there has not been a way of extending this approach to statistically relevant numbers of living cells and organisms. Herein, we design and fabricate a high throughput nanosensor array in a microfluidic channel that addresses this limitation, creating a Nanosensor Chemical Cytometry (NCC). An array of nIR fluorescent single walled carbon nanotube (SWNT) nanosensors is integrated along a microfluidic channel through which a population of flowing cells is guided. We show that one can utilize the flowing cell itself as highly informative Gaussian lenses projecting nIR emission profiles and extract rich information on a per cell basis at high throughput. This unique biophotonic waveguide allows for quantified cross-correlation of the biomolecular information with physical properties such as cellular diameter, refractive index (RI), and eccentricity and creates a label-free chemical cytometer for the measurement of cellular heterogeneity with unprecedented precision. As an example, the NCC can profile the immune response heterogeneities of distinct human monocyte populations at attomolar (10-18 moles) sensitivity in a completely non-destructive and real-time manner with a rate of ~100 cells/frame, highest range demonstrated to date for state of the art chemical cytometry. We demonstrate distinct H2O2 efflux heterogeneities between 330 and 624 attomole/cell·min with cell projected areas between 271 and 263 µm2, eccentricity values between 0.405 and 0.363 and RI values between 1.383 and 1.377 for non-activated and activated human monocytes, respectively. Hence, we show that our nanotechnology based biophotonic cytometer has significant potential and versatility to answer important questions and provide new insight in immunology, cell manufacturing and biopharmaceutical research.