Background: Anxiety and depression symptoms are common among cardiac patients. The Hospital Anxiety and Depression Scale (HADS) is frequently used to measure symptoms of anxiety and depression; however, no study on the validity and reliability of the scale in Danish cardiac patients has been done. The aim, therefore, was to evaluate the psychometric properties of HADS in a large sample of Danish patients with the four most common cardiac diagnoses: ischemic heart disease, arrhythmias, heart failure and heart valve disease. Methods: The DenHeart study was designed as a national cross-sectional survey including the HADS, SF-12 and HeartQoL and combined with data from national registers. Psychometric evaluation included analyses of floor and ceiling effects, structural validity using both exploratory and confirmatory factor analysis and hypotheses testing of convergent and divergent validity by relating the HADS scores to the SF-12 and HeartQoL. Internal consistency reliability was evaluated by Cronbach's alpha, and differential item functioning by gender was examined using ordinal logistic regression. Results: A total of 12,806 patients (response rate 51%) answered the HADS. Exploratory factor analysis supported the original two-factor structure of the HADS, while confirmatory factor analysis supported a three-factor structure consisting of the original depression subscale and two anxiety subscales as suggested in a previous study. There were floor effects on all items and ceiling effect on item 8. The hypotheses regarding convergent validity were confirmed but those regarding divergent validity for HADS-D were not. Internal consistency was good with a Cronbach's alpha of 0.87 for HADS-A and 0.82 for HADS-D. There were no indications of noticeable differential item functioning by gender for any items. Conclusions: The present study supported the evidence of convergent validity and high internal consistency for both HADS outcomes in a large sample of Danish patients with cardiac disease. There are, however, conflicting results regarding the factor structure of the scale consistent with previous research. Trial registration: ClinicalTrials.gov: NCT01926145.