A combined test of intestinal permeability using lactulose (L) and rhamnose (R), and absorptive function using xylose (X) and 3-O-methylglucose (G), was carried out at four, six, eight and 16 weeks of age in 22 healthy control and six gluten-sensitive Irish setter (IS) dogs fed a diet containing a controlled dose of gluten from weaning. Comparisons were made with two groups of 12 healthy control dogs of breeds other than IS, one fed the same diet as the setters and the other fed a gluten-free diet. Gluten-sensitive IS showed a rise in permeability (mean [SEM] urinary L/R) from 0.23 (0.07) at four weeks to 0.39 (0.05) at eight weeks, remaining at 0.36 (0.04) at 16 weeks. These results were significantly higher in gluten-sensitive than control IS at six, eight and 16 weeks, compatible with jejunal biopsy lesions characteristic of gluten-sensitive enteropathy demonstrated in affected dogs at 16 weeks. Urinary L/R ratios of control dogs of breeds other than IS peaked at six weeks 0.27 (0.02), and were significantly higher than those of control IS at six and eight weeks, demonstrating differences in permeability between Irish setter dogs and other breeds at this age. There were no significant differences in urinary X/G ratios at six, eight and 16 weeks of age between any of the groups of dogs challenged with gluten. Urinary L/R and X/G ratios were similar in the control dogs of breeds other than IS fed gluten-containing and gluten-free diets. These findings indicate that intestinal permeability testing of puppies during controlled oral gluten challenge provides a practical screening test for gluten sensitivity in Irish setter dogs at an early age.