Microfluidic Flow Cytometer Detection Platform Based on Spatially Modulated Fluorescence Emissions

We will present an optical detection technique that delivers high signal-to-noise discrimination without precision optics to enable a flow cytometer that can combine high performance, robustness, compactness and low cost. The enabling technique is termed “spatially modulated emission” and generates a time-dependent signal as a continuously fluorescing (bio-)particle traverses a optical transmission pattern. Correlating the detected signal with the known pattern achieves high discrimination of the particle signal from background noise. The analyte’s fluidic path, the optical excitation and detection volume need to be aligned for reliable particle detection. In conventional flow cytometry, the size of the excitation and detection volume is restricted approximately to the size of the particle. Our method uses a much larger excitation volume along the fluidic channel in order to increase the total flux of fluorescence light that originates from a particle while requiring minimal optical alignment. Despite the large excitation volume, the mask patterning enables a high spatial resolution in the micron range. This allows for detection and characterization of particles with a separation (in flow direction) comparable to the dimension of individual particles. In addition, the concept is intrinsically tolerant of background fluorescence originating from fluorescent components in solution, fluorescing components of the chamber and contaminants on the surface.

The fundamental advantages of the spatially modulated emission can be utilized in different ways. For a low-cost, robust, point-of-need absolute CD4+ and percentage CD4 counter for human blood, the detection technique has been extensive evaluated. By probing the same samples, we directly compared our system with a commercial instrument (BD FACSCount) and obtained excellent agreement for both absolute CD4 and percentage CD4. While for this application cost and size are crucial parameters we will also present our results on sample throughput, sheathflow-to-analyte ratio, sensitivity and multi-wavelength.