Optimized Approach for Microarray Scanning

摘要

Two common approaches to imaging fluorescent microarrays include CCD block scan imagers and laser/PMT-based scanners. CCD scanners afford high detector quantum efficiency, simultaneous illumination and detection of multiple pixels (parallelism) with concomitant opportunity to avoid dye saturation and difficult mechanical challenges -contributing to more reproducible measurements. CCD block scanners suffer from artifacts induced by out of focal-plane fluorescent particles - whose blurred image might not be analytically separable from target signal. Confocal scanners have excellent rejection of out-of-plane signals, and their small field of view allows for fine resolution with extremely high numerical apertures in the detection optics. Furthermore, they do not suffer from "stitching" artifacts commonly found in CCD systems that assemble a large image by tiling multiple blocks. Confocal systems scan continuously point by point, yet their design can be increasingly problematic as attempts are made to combine rapid scanning speeds, tight resolution, and large numerical apertures, where, for example, limitations in the depth of field can be especially worrisome. This paper explores some important principles governing SNR and susceptibility to artifacts for these strategies. Consideration is given to high dynamic range and high-sensitivity scanning approaches based upon CCD's that reap the aforementioned benefits of PMT-based confocal scanners.