Groundtruth approach to accurate quantitation of fluorescence microarrays

Laura Mascio Kegelmeyer; Lisa Tomascik-Cheeseman; Melinda S. Burnett; Paul van Hummelen; Andrew J. Wyrobek

doi:10.1117/12.427998

4 June 2001 Groundtruth approach to accurate quantitation of fluorescence microarrays

Laura Mascio Kegelmeyer, Lisa Tomascik-Cheeseman, Melinda S. Burnett, Paul van Hummelen, Andrew J. Wyrobek

Proceedings Volume 4266, Microarrays: Optical Technologies and Informatics; (2001) https://doi.org/10.1117/12.427998
Event: BiOS 2001 The International Symposium on Biomedical Optics, 2001, San Jose, CA, United States

Abstract

To more accurately measure fluorescent signals from microarrays, we calibrated our acquisition and analysis systems by using groundtruth samples comprised of known quantities of red and green gene-specific DNA probes hybridized to cDNA targets. We imaged the slides with a full-field, white light CCD imager and analyzed them with our custom analysis software. Here we compare, for multiple genes, results obtained with and without preprocessing (alignment, color crosstalk compensation, dark field subtraction, and integration time). We also evaluate the accuracy of various image processing and analysis techniques (background subtraction, segmentation, quantitation and normalization). This methodology calibrates and validates our system for accurate quantitative measurement of microarrays. Specifically, we show that preprocessing the images produces results substantially closer to the known groundtruth for these samples.

Citation Download Citation

Laura Mascio Kegelmeyer, Lisa Tomascik-Cheeseman, Melinda S. Burnett, Paul van Hummelen, and Andrew J. Wyrobek "Groundtruth approach to accurate quantitation of fluorescence microarrays", Proc. SPIE 4266, Microarrays: Optical Technologies and Informatics, (4 June 2001); https://doi.org/10.1117/12.427998

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