A standard for the assessment of lactose malabsorption does not exist. As measured by lactose tolerance tests, insufficient increase in blood glucose or increased breath hydrogen (H2) excretion after lactose ingestion is regarded as pathological. In this study, we have tried to elucidate the relationship between lactose tolerance test results and symptoms after a lactose challenge. This relationship might be an indicator for the validity of the test.In a prospective study, 309 consecutive patients with suspected lactose malabsorption underwent a lactose tolerance test. After consumption of 50 g of lactose, blood glucose and breath H2 concentrations were measured. During the test (240 min), the severity of bloating, flatulence, abdominal distention, and diarrhea were semiquantitatively scored as 0, 1, or 2. The individual sum of these four scores was calculated and denoted as the total symptom score (TSS). All subjects were classified according to their TSS to compare symptoms with peak breath-H2 concentration and change in blood glucose concentration, respectively.The glucose and breath H2 response were pathological in 51.1 and 39.5% of cases, respectively. A stepwise increase in TSS of 1 point was associated with a significant increase (p < 0.05) in mean peak H2 concentration. However, a significantly lower glucose increment compared with patients with a TSS of 0 was found only in patients with a TSS of 2 or 4. The mean symptom score differed significantly between the positive and negative breath tests (p < 0.001), but did not differ between the positive and negative glucose response results.This study shows that GI symptoms after a lactose challenge are strongly associated with the amount of H2 excretion. The relationship between the increase in glucose concentration and symptoms after a lactose load is less evident. Thus, the H2 breath test seems to be superior to the measurement of blood glucose increment as a diagnostic tool in lactose malabsorption, although the true predictive value of this test only can be determined after a period of dietary treatment.