Flow cytometer system for single-shot biosensing based on whispering gallery modes of fluorescent microspheres

We report an innovative label-free biosensor based on statistical analysis of several whispering gallery modes spectral shifts in polystyrene fluorescent microspheres using a custom microflow cytometer. Whispering gallery modes analysis enables detection of nanometer-sized analytes showing promising possibilities for virus, bacteria and molecular detection. To demonstrate this, fluorophore-doped microspheres of the appropriate size parameter are mixed in an aqueous solution. Then, a syringe pump pushes the solution through a fiber optic flow cell where a laser beam illuminates the analysis area to excite the microspheres and their fluorescence is collected. This device provides a low-cost and user friendly solution that could enhance spectrum acquisition rates up to 5 spectra per second thanks to the considerable amount of microspheres flowing through the excitation area per unit time. Finally, the fluorescence spectra are statistically investigated using an instantaneous measurement of apparent refractive index algorithm to determine a reliable value for the refractive index of the environment since the exact radius of the microsphere scanned is unknown. This refractive index becomes an effective value for the local perturbation caused by inhomogeneities on the microsphere surface and hence, determines whether or not inhomogeneities, such as bacteria, are adsorbed by comparing to a control sample. Combining a flow cell with our detection algorithm, we reduce the period of a 50 microspheres experiment from 161 minutes to 14 minutes when the flow rate is 2000 µl/h and the microsphere concentration is 5 µsphere/µl.