Objective: Validate the acromegaly quality of life (AcroQoL) questionnaire as a disease-generated questionnaire, which analyses physical and psychological domains, the latter subdivided into appearance and personal relationship sub-scales, to evaluate health-related quality of life (HRQoL) in acromegaly. Design: Prospective, observational multicenter study. Methods: One hundred and six patients with acromegaly, 42 with active disease studied basally and 6 months after treatment ('sensitivity to change' group), and 64 with treated, stable disease, studied twice within 1 month ('reliability' group) were included. As controls, a reference Spanish population (n=12 245 for the EuroQoL questionnaire) and 157 obese patients (body mass index>30 kg/m2) were studied basally. Socio-demographic data, clinical activity, co-morbidity, GH, IGF-I, and HRQoL (overall perception of health state, EuroQoL and AcroQoL in the obese controls and acromegalic patients) were evaluated. Results: Globally, AcroQoL scored worse in the 'sensitivity to change' group than in the 'reliability' group (56±20 vs 65±18, P<0.05), but did not discriminate between patients and obese controls. The psychological domain was worse in the 'sensitivity to change' group than obese controls (P<0.05). Appearance was the most affected sub-scale in acromegaly and significantly worse than in obese controls. The sub-scale personal relationships of AcroQoL were less affected in the 'reliability' group than in obese controls (P<0.05). Patients with acromegaly and obese controls showed more problems on the EuroQoL than general Spanish population. Significant correlations were observed globally and for each dimension between AcroQoL and the generic questionnaires. On re-testing, no change was observed in the 'reliability' group in any questionnaire, demonstrating good test-re-test reliability. In the 'sensitivity to change' group after 6 months of treatment, there was improvement in the generic questionnaires and in AcroQoL score (P<0.01). Internal consistency of AcroQoL was good (Cronbach's α>0.7). No correlation between AcroQoL and GH or IGF-I was observed. Conclusion: AcroQoL questionnaire is a valid tool for the assessment of HRQoL in clinical practice in patients with acromegaly. © 2006 Society of the European Journal of Endocrinology.