Fluorescence array detector for large‐field quantitative fluorescence cytometry

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Abstract
A prototypic fluorescence array detector (FAD) has been designed and constructed which is capable of quantifying single‐cell fluorescence emissions from a statistically significant population of cell‐sized objects (over 103) on a solid substrate. The system is comprised of a cryogenically cooled CCD, 50 mW air‐cooled argon ion laser, and optics that image a large (1 × 1 cm) field at 1:1 (no magnification). The CCD is effectively treated as a two‐dimensional array of 2.7 × 105 independent 20 × 20 μm photodetectors, with each cell‐sized object imaged across only a few CCD pixels. Algorithms have been developed for focusing, image segmentation, shading correction, and noise rejection; performance data for the FAD with fluorescent calibration beads are presented. The FAD is a simple alternative to microscope‐based imaging cytometry, allowing large‐field imaging without a scanning stage.