  Despite the success and popularity of oligonucleotide arrays as a
high-throughput technique for measuring mRNA expression levels, quantitative
calibration studies have until now been limited. The main reason is that
suitable data was not available. However, calibration data recently produced by
Affymetrix now permits detailed studies of the intensity dependent sensitivity.
Given a certain transcript concentration, it is of particular interest to know
whether current analysis methods are capable of detecting differential
expression ratios of 2 or higher. Using the calibration data, we demonstrate
that while current techniques are capable of detecting changes in the low to
mid concentration range, the situation is noticeably worse for high
concentrations. In this regime, expression changes as large as 4 fold are
severely biased, and changes of 2 are often undetectable. Such effects are
mainly the consequence of the sequence specific binding properties of probes,
and not the result of optical saturation in the fluorescence measurements.
GeneChips are manufactured such that each transcript is probed by a set of
sequences with a wide affinity range. We show that this property can be used to
design a method capable of reducing the high intensity bias. The idea behind
our methods is to transfers the weight of a measurement to a subset of probes
with optimal linear response at a given concentration, which can be achieved
using local embedding techniques.
