To establish a high performance liquid chromatography-tandem hybrid triple-quadrupole linear ion trap mass-spectrometry method for qualitative and quantitative determination of etomidate in blood and hair. Deuterium cocaine was selected as the internal standard, and blood sample was extracted by acetonitrile (V/V, 1/6), then centrifuged at high speed; hair sample (about 20 mg) was ground by a ball mill, then extracted by methanol and passed through the organic membrane. Using 0.1% (V/V) formic acid aqueous solution and 0.1% (V/V) formic acid acetonitrile as mobile phase, the analyte was separated and analyzed by an ACQUITY UPLC^®C18 (2.1 mm×100 mm×1.7 μm) column. Electrospray positive multiple reaction monitoring/information-dependent acquisition/enhanced product ion scanning (MRM-IDA-EPI) and secondary library retrieval were used for analysis. The results showed that etomidate in blood had a good linear relationship in the range of 1.0 to 100.0 ng/mL (r > 0.995). The linear relationship of etomidate in hair was good in the range of 0.05 to 5.0 ng/mg (r > 0.995). The detection limits (S/N ≥ 3) for blood and hair were 0.2 ng/mL and 0.002 ng/mg, and the quantitative limits (S/N ≥ 10) were 0.5 ng/mL and 0.005 ng/mg respectively. The recoveries of 1.0, 10.0, 100.0 ng/mL in spiked blood were 97.1% to 103.4%, and 0.5, 2.5, 5.0 ng/mg in spiked hair were 84.0% to 99.8%. The relative standard deviation was less than 15%. This method can be applied to the qualitative and quantitative analysis of etomidate in blood and hair in the juridical practice.