To investigate the effect of systemic chloroquine/hydroxychloroquine (CQ/HCQ) on outer retinal health using quantitative fundus autofluorescence (QAF) imaging. Methods: For this prospective, cross-sectional study, 44 CQ/HCQ patients and 25 age-matched controls underwent multimodal retinal imaging including QAF (488 nm) and spectral-domain optical coherence tomography (SD-OCT) in addition to the recommended CQ/HCQ screening procedures. Custom written FIJI plugins enabled detailed QAF analysis and correlation with retinal thickness and comparison to the healthy controls. Results: Out of 44 patients, 29 (mean age 43.5 ± 12.2, range 22-59 years) exposed to CQ/HCQ (mean cumulative dose 724.2 ± 610.4 g, median 608.0 g, range 18.6-2171.0 g) met eligibility criteria. Four of these 29 patients had bull's-eye maculopathy (BEM). Mean QAF values were significantly higher in CQ/HCQ patients than in healthy controls. QAF increase started early after treatment onset, remained high even years after treatment cessation, and was not accompanied by pathologies in the other screening methods, including retinal thicknesses (except in BEM patients). Conclusions: QAF might be a useful tool in retinal imaging and in verifying systemic CQ/HCQ intake. The early onset and preserved high levels of QAF parallel findings of CQ deposition in the retina in animal models. Whether QAF can be used as a screening tool to detect early CQ/HCQ related maculopathy is the subject of long-term ongoing studies. Translation Relevance: Experimental QAF imaging in systemic CQ/HCQ therapy monitoring might be a useful tool to indicate the drug or its metabolites and to detect metabolic retinal changes.