Purpose: This study assesses the repeatability of quantitative autofluorescence (qAF) in a multicenter setting and evaluates qAF as the end point for clinical trials in recessive Stargardt disease... Show morePurpose: This study assesses the repeatability of quantitative autofluorescence (qAF) in a multicenter setting and evaluates qAF as the end point for clinical trials in recessive Stargardt disease 1 (STGD1).Methods: A total of 102 patients with STGD1 underwent qAF imaging as part of the Stargardt Remofuscin Treatment Trial (STARTT; EudraCT No. 2018-001496-20). For 166 eyes, we obtained qAF imaging at 2 visits, with 2 recordings per visit. The qAF8 values were independently determined by the study site and a central reading center. Intra-and inter-visit reproducibility, as well as interobserver (study site versus reading center) reproducibility were obtained using intraclass correlation (ICC), one-sample t-test, and Bland-Altman coefficient of repeatability.Results: The qAF repeatability was & PLUSMN; 26.1% for intra-visit, & PLUSMN; 40.5% for inter-visit, and & PLUSMN; 20.2% for the interobserver reproducibility measures. Intra-visit repeatability was good to excellent for all sites (ICC of 0.88-0.96). Variability between visits was higher with an overall ICC of 0.76 (0.69-0.81). We observed no significant difference in qAF values across sites between visits (7.06 & PLUSMN; 93.33, P = 0.238).Conclusions: Real-life test-retest variability of qAF is higher in this set of data than previ-ously reported in single center settings. With improved operator training and by select-ing the better of two recordings for evaluation, qAF serves as a useful method for assess-ing changes in autofluorescence signal.Translational Relevance: The qAF can be adopted as a clinical trial end point, but steps to counterbalance variability should be considered. Show less