Accurate quantitation of target genes depends on correct normalization. Use of genes with variable tissue transcription (GAPDH) is problematic, particularly in clinical samples which are derived from different tissue sources. Using a large-scale gene database (Affymetrix U133A) dataset of 36 gastrointestinal tumors and normal tissues, we identified 8 candidate reference genes and established expression levels by real-time RT-PCR in an independent dataset (n=42). A geometric averaging method (geNorm) identified ALG9, TFCP2 and ZNF410, as the most robustly expressed control genes. Examination of raw CT values demonstrated these genes were tightly correlated between themselves (R²>0.86, p<0.0001) with low variability (CV <12.7%) and high inter-assay reproducibility (r=0.93, p=0.001). In comparison, the alternative control gene, GAPDH, exhibited the highest variability (CV=18.1%), was significant differently expressed between tissue types (p=0.05), was poorly correlated with the 3 reference genes (R²<0.4) and was considered the least stable gene. To illustrate the importance of correct normalization, the target gene, MTA1, was significantly over-expressed (p=0.0006) in primary GI NET samples (versus normal GI samples) when normalized by geNorm_ATZ but not when normalized using GAPDH. The geNorm_ATZ approach was, in addition, applicable to adenocarcinomas; MTA1 was over-expressed (p<0.04) in malignant colon, pancreas and breast tumors compared to normal tissues. We provide a robust basis for the establishment of a reference gene set using GeneChip data and provide evidence for the utility of normalizing a malignancy-associated gene (MTA1) using novel reference genes and the geNorm approach in gastrointestinal NETs as well as in adenocarcinomas and breast tumors.