Measurements in flow: improving cytometry and flow metrology in optofluidic devices

G. Cooksey
National Institute of Standards and Technology,
United States

Keywords: microfluidic, measurements, optofluidic devices


Microfluidic components are critical to many biological and chemical measurement systems. High surface-to-volume ratios and other unique physical properties can be exploited to achieve various useful conditions, but better measurements are needed to determine uncertainties in microscale environments. Microfluidic platforms with integrated optical components offer great promise as robust platforms to interrogate the nature and composition of microflows. We have recently demonstrated an optofluidic measurement system that improves liquid flow measurements more than 10-fold, achieving 5 % uncertainty at 1 nL/min. We are also using these systems to address measurement challenges in flow cytometry. Though flow cytometers can make hundreds of single-cell measurements per second, per-particle measurement uncertainty is difficult to determine because it is confounded by variability in the sample population and flow conditions. To better investigate the measurement process, we have developed a new kind of cytometer that repeats interrogation of objects at multiple points along a flow path. Critically, the independent measurement regions enable direct quantification of uncertainties associated with effects arising from operating conditions. The system was used to determine per-particle uncertainty based on sheath focusing, which enabled study and utilization of inertial effects to reduce measurement variation. We achieved less than 2 % measurement variation per particle with greater than 99.9 % tracking accuracy. Because the system captures a complete profile of objects as they pass through each measurement region, we can further improve recognition and separation of coincidence events (or doublets, which can’t be done conventionally) and study the shape of objects under different flow conditions. More recent improvements in optical components have demonstrated limits of detection in our microcytometer are below conventional flow cytometers. Overall, our systems are addressing important measurement challenges in cytometry, such as separating true population heterogeneity from measurement variation and quantitative classification of sample composition, including rare-event detection.