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These cytokines have been implicated in the pathogenesis of autoimmunity, most notably systemic lupus erythematosus (SLE), diabetes mellitus, and dermatomyositis, as well as monogenic type I interferonopathies. Despite a fundamental role in health and disease, the direct quantification of type I IFNs has been challenging. Using single-molecule array (Simoa) digital ELISA technology, we recorded attomolar concentrations of IFN\u03b1 in healthy donors, viral infection, and complex and monogenic interferonopathies. IFN\u03b1 protein correlated well with functional activity and IFN-stimulated gene expression. High circulating IFN\u03b1 levels were associated with increased clinical severity in SLE patients, and a study of the cellular source of IFN\u03b1 protein indicated disease-specific mechanisms. 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