Measurements of plasma free metanephrines (MNs), including MN and normetanephrine, provide high sensitivity and specificity for the diagnosis of pheochromocytoma and paraganglioma (PPGL). 3-Methoxytyramine (3-MT) and chromogranin A (CgA) may allow the detection of dopamine-producing or biochemically silent PPGL. The aim of this study was to evaluate whether measurements of plasma 3-MT or CgA as a supplement of plasma MNs offer a better diagnostic strategy for initial testing of PPGL.We enroled 125 patients who underwent surgery from 2015 to 2016 for our study and identified 33 patients with PPGL and 92 patients with non-PPGL masses.The levels of plasma free MNs and 3-MT were measured for all 125 patients using liquid chromatography-tandem mass spectrometry. Plasma CgA concentrations were determined using a radioimmunoassay. To evaluate the diagnostic performance of plasma free MNs, 3-MT and CgA, sensitivity and specificity were determined, and receiver operating characteristic curves were constructed.We found that combining 3-MT and MNs increased the diagnostic sensitivity from 93.9% (95% confidence interval [CI]: 78.4%-98.9%) to 97.0% (95% CI: 82.5%-99.8%). In contrast, addition of plasma CgA test reduced the diagnostic specificity significantly from 91.3% (95% CI: 83.1%-95.9%) to 75.0% (95% CI: 64.7%-83.2%).Here, we demonstrated that 3-MT represents a valuable supplementary test to plasma MNs, which can further enhance the sensitivity of the assay, while plasma CgA added no additional diagnostic value to MNs due to the lowered diagnostic specificity.© 2021 John Wiley & Sons Ltd.