Background Accurate quantification of iron load is of importance in tailoring the therapy for patients with iron load disease. Magnetic resonance (MR) imaging is commonly used to assess iron load in different organs by calculating the T2* value. Estimation of precision and uncertainty of the obtained T2* value may be useful for determining changes in iron load between follow-up scans and for titration of treatment. Therefore, the purpose of this study was to develop and validate a new T2* analysis method, providing certainty estimates for quantification of iron load in heart and liver. Methods The proposed method is ar egion of interest (ROI)based algorithm which changes curve-fitting procedure based on the results from an initial T2* estimate using a 3-parameter, noise-corrected model. The certainty estimate, presented as the T2* value confidence interval, is derived from multiple sub-regions within the selected ROI. Except for ROI delineation, the proposed method does not require any user interaction. Phantom imaging was performed on a 1.5T system, with two clinically available multi-echo gradient-recalled echo sequences for cardiac and liver imaging. Phantoms were constructed to cover the clinically important range of T2*. A T2* single-echo gradient-echo sequence with TR set to 6 × T1 and a 50° flip angle was used as reference standard. Computer simulations were performed to assess accuracy and precision from 2 000 repetitions at SNR=15. Inter- and intra-observer variability was obtained in patients (n=22) by one experienced and one inexperienced observer.