To identify and characterize genes, the products of which play a role in pancreatic adenocarcinoma, we constructed a complementary DNA (cDNA) library using mRNA from the pancreatic adenocarcinoma cell line HPAF, grown as a nude mouse tumor. Through differential screening, we identified a cDNA clone, pII5B, that is homologous to an mRNA expressed at significantly higher levels in HPAF cells than in normal human pancreas. The pII5B cDNA was homologous to the 3'-untranslated region of glyceraldehyde-3-phosphate dehydrogenase (GAPDH, EC 1.2.1.12)mRNA. Partial sequencing of several HPAF tumor GAPDH cDNA clones revealed no significant differences from previously published GAPDH cDNA sequences. Increased levels of GAPDH mRNA, relative to actin mRNA levels, were found in six pancreatic adenocarcinoma cell lines and two nude mouse tumors, when compared to normal pancreas. Enolase and glucose transporter mRNA levels were also increased in HPAF cells and nude mouse tumor, suggesting a general increase in expression of genes associated with glycolysis in pancreatic adenocarcinoma. Levels of GAPDH protein were elevated in nude mouse tumors and fresh human pancreatic adenocarcinomas compared to normal pancreas. High GAPDH levels may be characteristic of human adenocarcinomas, since colon adenocarcinomas also exhibited high levels of GAPDH compared to normal colon.