Highly Sensitive Cy3 Fluorescence Detection Method for Microarray Based on CCD and LED

Microarrays play a crucial role in detecting diverse biological assays, supported by high-throughput fluorescent scanners that stem from laser confocal scanning mechanisms or CCD wide-field imaging technology. Nonetheless, each scanner exhibits significant limitations. To address these drawbacks, this study proposes a novel design for detecting Cy3 fluorescence microarrays using a cooled CCD and high-power LED, which demonstrates excellent performance to overcome the high complexity of the laser confocal technique and the low efficiency of high-pressure gas lamps in traditional CCD scanner by ExviewHAD CCD of ICX694AL and single-core powerful LED of PT54G. The experimental results, including comparative testing of MGMED BAC microarrays using laser confocal scanning and cooled CCD reading, reveal that the novel CCD scanner achieves a detection sensitivity of 1.12 fluor/μm ² , a reading resolution of 10.0μm, and a measuring repeatability indicator R_v ≤ 2.33%. Additionally, to mitigate no-linear response in quantitative detection of Cy3 fluorophores, the calibration curve is analyzed by the least squares approximation (LSA), and the LSA fitting of N=3 in the CCD linearity region yields a maximum relative error of only 2.35%. These exceptional performance metrics indicate that this newly developed CCD wide-field scanner integrated with a high-power LED, exhibits excellent capability in detecting Cy3 microarrays.