Figure 2.

The calibration model for frequency normalization. Eleven control transcripts are spiked into the hybridization solution at known concentrations, and the absolute difference (AD) measurements for these controls are plotted against their known frequencies. P and A represent Present and Absent calls, respectively, from Affymetrix GeneChip software. Hybridization response in average difference (AD) is approximately proportional to transcript abundance. The solid fitted line is a linear model with intercept zero, which is used to calculate frequencies for all other transcripts on the array. The vertical line at 4.5 ppm represents the calculated limit of detection for this particular array; frequencies below that level are damped to avoid attributing significance to expression differences caused by assay noise.