We conducted a systematic review of the accuracy of fundus autofluorescence (FAF) imaging for diagnosing and monitoring retinal conditions. Searches in November 2014 identified English language references. Sources included MEDLINE, EMBASE, the Cochrane Library, Web of Science, and MEDION databases; reference lists of retrieved studies; and internet pages of relevant organisations, meetings, and trial registries. For inclusion, studies had to report FAF imaging accuracy quantitatively. Studies were critically appraised using QUADAS risk of bias criteria. Two reviewers conducted all review steps. From 2240 unique references identified, eight primary research studies met the inclusion criteria. These investigated diagnostic accuracy of FAF imaging for choroidal neovascularisation (one study), reticular pseudodrusen (three studies), cystoid macular oedema (two studies), and diabetic macular oedema (two studies). Diagnostic sensitivity of FAF imaging ranged from 32 to 100% and specificity from 34 to 100%. However, owing to methodological limitations, including high and/or unclear risks of bias, none of these studies provides conclusive evidence of the diagnostic accuracy of FAF imaging. Study heterogeneity precluded meta-analysis. In most studies, the patient spectrum was not reflective of those who would present in clinical practice and no studies adequately reported whether FAF images were interpreted consistently. No studies of monitoring accuracy were identified. An update in October 2016, based on MEDLINE and internet searches, identified four new studies but did not alter our conclusions. Robust quantitative evidence on the accuracy of FAF imaging and how FAF images are interpreted is lacking. We provide recommendations to address this.