Incretin hormones importantly enhance postprandial insulin secretion but are rapidly degraded to inactive metabolites by ubiquitous dipeptidyl peptidase IV. The concentrations of the intact biologically active hormones remain largely unknown. Using newly developed assays for intact glucagon-like peptide (GLP)-1 and glucose-dependent insulinotropic polypeptide (GIP), we measured plasma concentrations after a mixed breakfast meal (566 kcal) in 12 type 2 diabetic patients (age 57 years [range 49–67], BMI 31 kg/m² [27–38], and HbA_1c 9.2% [7.0–12.5]) and 12 matched healthy subjects. The patients had fasting hyperglycemia (10.7 mmol/l [8.0–14.8]) increasing to 14.6 mmol/l (11.5–21.5) 75 min after meal ingestion. Fasting levels of insulin and C-peptide were similar to those of the healthy subjects, but the postprandial responses were reduced and delayed. Fasting levels and meal responses were similar between patients and healthy subjects for total GIP (intact + metabolite) as well as intact GIP, except for a small decrease in the patients at 120 min; integrated areas for intact hormone (area under the curve [AUC]_INT) averaged 52 ± 4% (for patients) versus 56 ± 3% (for control subjects) of total hormone AUC (AUC_TOT). AUC_INT for GLP-1 averaged 48 ± 2% (for patients) versus 51 ± 5% (for control subjects) of AUC_TOT. AUC_TOT for GLP-1 as well as AUC_INT tended to be reduced in the patients (P = 0.2 and 0.07, respectively); but the profile of the intact GLP-1 response was characterized by a small early rise (30–45 min) and a significantly reduced late phase (75–150 min) (P < 0.02). The measurement of intact incretin hormones revealed that total as well as intact GIP responses were minimally decreased in patients with type 2 diabetes, whereas the late intact GLP-1 response was strongly reduced, supporting the hypothesis that an impaired function of GLP-1 as a transmitter in the enteroinsular axis contributes to the inappropriate insulin secretion in type 2 diabetes.