Few self-report measures of medication adherence have been rigorously developed and validated against electronic drug monitoring (EDM). Assess the validity of the 3-item self-report scale by comparing it with a contemporaneous EDM measure. We conducted an observational study in which adherence assessments were done monthly for up to 4 months for 81 patients with HIV who were taking antiretroviral medications. We report results for both HIV antiretroviral medications, and also for other, non-HIV-related medications. Raw and calibrated self-report adherence measures, electronic drug monitoring adherence measures, and sociodemographic variables. The mean age of patients was 46 years, 37 % were female, 49 % had some education beyond high school, 22 % were Black, and 22 % were Hispanic. Cronbach's alphas for the 3-item scale for HIV and non-HIV medications were 0.83 and 0.87, respectively. The mean differences (raw/uncalibrated self-report scale minus EDM) for HIV and non-HIV medications were 7.5 and 5.2 points on a 100-point scale (p < 0.05 for both). Pearson correlation coefficients between the calibrated 3-item scale and the EDM for HIV and non-HIV medications were 0.47 and 0.59, respectively. The c-statistics for the ROC curves for the calibrated scale, using cut-offs of 0.8 and 0.9 for the EDM gold standard measure to define non-adherence, were between 0.74 and 0.76 for HIV and non-HIV medications. This 3-item adherence self-report scale showed good psychometric characteristics and good construct validity when compared with an EDM standard, for both HIV and non-HIV medications. In clinical care it can be a useful first-stage screener for non-adherence. In clinical research and quality improvement settings it can be a useful tool when more complex and expensive methods such as EDM or pharmacy claims are impractical or unavailable.