Array imaging and data analysis

Microarray slides stained with RLS particles were imaged at a resolution of 10 mm with a 16 bit CCD camera-based scanner (Invitrogen) and images analyzed with ArrayVision, version 8.0 (Imaging Research, St. Catharine's, Canada). Median-trimmed mean signal values for each spot on a slide were imported into EXCEL. For each slide, standard curves for each analyte were generated by four-parameter logistic fitting. Unknown sample concentrations were calculated using the corresponding signal values, the curve fitting parameters, and the dilution factors. An average concentration (derived from the three replicate spots) was calculated for each of the dilutions. To obtain a single concentration value, the program automatically chose the lowest dilution that gave a signal in the dynamic range ofthe assay. We performed one four-slide experiment for each ofthe two antibody array panels. For each ofthe four HUPO reference specimen preparations, three aliquots ofthe Asian-American, African-American, and Chinese samples and one aliquot of the NIBSC citrate-plasma reference sample were incubated on the same slide and measured against the same set of standard curves.

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