Fluorescence array detector for large-field quantitative fluorescence cytometry

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 10(3)) 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 x 1 cm) field at 1:1 (no magnification). The CCD is effectively treated as a two-dimensional array of 2.7 x 10(5) independent 20 x 20 microns 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.