This study is aimed to establish whether the skin preparation using ethanol (EA)-containing skin antiseptics causes EA contamination through blood collection. Venous blood was collected from 40 healthy volunteers according to the national guidelines for blood sampling, with four sequential procedures as follows: 1) collecting immediately (within 5 s) after cleaning the skin with each packaging type of EA-containing pad, 2) collecting 1 minute after cleaning the skin with each packaging type of EA-containing pad, 3) collecting immediately (within 5 s) after cleaning the skin with a traditional cleaning method (with a thoroughly EA-impregnated pad, and 4) collecting 1 minute after cleaning the skin with a traditional cleaning method. Each sequential procedure was performed with and without the EA-containing pad used for skin cleaning pressed against the puncture site on their right and left arms at the time the needle was withdrawn, respectively. The collected specimens were subjected to determination of EA by using head space gas chromatography-mass spectrometry.　In every 80 blood specimen obtained from 40 participants, EA was undetectable (<0.001 mg/mL). This study demonstrates that disinfection using ethanol-containing skin antiseptics is unlikely to cause ethanol contamination through blood collection regardless of skin preparation technique according to the guidelines for blood sampling.