Gene expression can be quantitatively analyzed by hybridizing fluor-tagged mRNA to targets on a cDNA microarray. Comparison of gene expression levels arising from cohybridized samples is achieved by taking ratios of average expression levels for individual genes. A novel method of image segmentation is provided to identify cDNA target sites and a hypothesis test and confidence interval is developed to quantify the significance of observed differences in expression ratios. In particular, the probability density of the ratio and the maximum-likelihood estimator for the distribution are derived, and an iterative procedure for signal calibration is developed.