FLUORESCENCE ARRAY DETECTOR FOR LARGE-FIELD QUANTITATIVE FLUORESCENCECYTOMETRY

A prototypic fluorescence array detector (FAD) has been designed and c onstructed which is capable of quantifying single-cell fluorescence em issions from a statistically significant population of cell-sized obje cts (over 10(3))on a solid substrate. The system is comprised of a cry ogenically cooled CCD, 50 mW air-cooled argon ion laser, and optics th at image a large (1 x 1 cm) field at 1:1 (no magnification). The CCD i s effectively treated as a two-dimensional array of 2.7 x 10(5) indepe ndent 20 x 20 mu m photodetectors, with each cell-sized object imaged across only a few CCD pixels. Algorithms have been developed for focus ing, image segmentation, shading correction, and noise rejection; perf ormance data for the FAD with fluorescent calibration beads are presen ted. The FAD is a simple alternative to microscope-based imaging cytom etry, allowing large-field imaging without a scanning stage. (C) 1994 Wiley-Liss, Inc.