  This paper describes and compares two methods for estimating the variance
function associated with iTRAQ (isobaric tag for relative and absolute
quantitation) isotopic labeling in quantitative mass spectrometry based
proteomics. Measurements generated by the mass spectrometer are proportional to
the concentration of peptides present in the biological sample. However, the
iTRAQ reporter signals are subject to errors that depend on the peptide
amounts. The variance function of the errors is therefore an essential
parameter for evaluating the results, but estimating it is complicated, as the
number of nuisance parameters increases with sample size while the number of
replicates for each peptide remains small. Two experiments that were conducted
with the sole goal of estimating the variance function and its stability over
time are analyzed, and the resulting estimated variance function is used to
analyze an experiment targeting aberrant signaling cascades in cells harboring
distinct oncogenic mutations. Methods for constructing conservative $p$-values
and confidence intervals are discussed.
