Objective To establish the normal reference range of serum tumor markets for health people in Shanghai to provide the basis for the clinical application. Method 604 healthy human serum which were collected from March 2012 to April 2013 have been tested by Flow Fluorescence Immunoassay for the concentration of AFP, CA125, CYFRA21-1, CEA, NSE, and CA19-9. Results The AFP reference range was <10.32ng/mL in male and <8.12ng/mLin female; CA125 reference range was 27.02U/mL; CEA reference range was <4.3ng/mL in male and <3.74ng/mLin female; NSE reference range was <11.64ng/mL in female with age less than 45years, <15.20ng/mL in female with age more than 45years, <18.47ng/mL in male wit age less than 45years and <20.04ng/mL with age more than 45 years; CA19-9 reference range was < 20.73U/mL; and tPSA reference range was <1.48ng/mL.Conclusion The preliminarily reference range of AFP, CA125, CYFRA21-1, CEA, NSE, CA19-9 for healthy people in Shanghai are established.