Gene regulation at the posttranscriptional level is often mediated by trans-acting factors binding the 3' untranslated region (3' UTR) of messenger RNAs (mRNAs). Alternative mRNA isoforms differing only in their 3' UTR can thus be differentially regulated, and it has been recently shown that this mechanism is indeed used by the cell to alter gene regulation effected by microRNAs and RNA-binding proteins, especially in highly proliferating contexts. Here we describe a computational method to analyze alternative 3' UTR isoforms in gene expression profiling datasets obtained with Affymetrix 3' IVT microarrays. The approach we describe allows the analysis of 3' UTR isoform usage in thousands of publicly available gene expression datasets, including many retrospective studies of cancer patients equipped with clinical data.