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. 2012 Jun 8;2:445. doi: 10.1038/srep00445

Figure 6. Flow cytometry with the HDLS.

Figure 6

(a) Comparison in interrogation method between the conventional flow cytometer and HDLS-based flow cytometer. While conventional flow cytometers cannot resolve multiple cells (i.e., doublets, triplets, and clustered cells) within the interrogation beam due to the lack of spatial resolution, spatial metrics provided by the HDLS can be used to identify these events and hence reduce false positive rate and statistical error yet with high throughput. (b) Experimental apparatus for the HDLS-based flow cytometry. The microfluidic channel focuses and orders randomly distributed cells into a single stream with inertial lift forces (Supplementary Information). HDLS pulses are then focused onto the single stream of fast-flowing cells for forward-scattering measurements. Scattered light from the target is collected by an objective lens and detected by a single-pixel photodetector. To ensure stability, the microfluidic device was fabricated in thermoset polyester (TPE) using standard replica molding methods (Supplementary Information). The volumetric flow rate of cells was controlled by a syringe pump, and a uniform velocity of 1 m/s was achieved by positioning cells precisely in a velocity gradient using inertial focusing. (c) Comparison of size-based histograms obtained with the conventional flow cytometer (left) and the HDLS-based flow cytometer (right) using identical samples of white blood cells and MCF7 breast cancer cells (measured separately). The figure clearly indicates our method's ability to differentiate MCF7 breast cancer cells from white blood cells as it can identify multiple white blood cells as false positive events. (d) Receiver operating characteristic (ROC) curve analysis showing that flow cytometry with the HDLS achieves lower false positive rate than flow cytometry without it, yet with high sensitivity and high throughput (Supplementary Information). This is made possible by the complete spectrum in the spatial frequency domain provided by the HDLS's ultrafast laser-scanning capability.