Flow-stream waveguide for collection of perpendicular light scatter in flow cytometry.

We report a new physical configuration for the detection of perpendicular light scatter or fluorescence in flow cytometry when using a fluid stream in air. This configuration increases the signal-to-noise ratio and narrows the coefficient of variation for uniformly sized latex spheres when compared to using a microscope objective to collect such light. The new technique views the scattered light that is trapped within the optical waveguide that is naturally formed by the flow stream in air. One efficient and simple way to detect the light trapped within this optical waveguide is to place one end of a fiber optic, with a conically polished tip and sufficiently large-core diameter, directly into the flow stream and to place an optical detector at the fiber's other end. For perpendicular light scatter, the flow-stream waveguide achieves high collection efficiency (NA = 0.88) as well as high efficiency of optical through put due to lack of surfaces between the light scatterers and the detector. We obtained 10-fold higher signals with this technique than with a long-working-distance microscope objective. The flow-stream waveguide is also much easier to align than traditional microscope-lens-based systems.