Objective To establish the reference intervals of serum osteocalcin (OCN), C-terminal cross-linking telopeptide of type Ⅰ collagen (β-CTx) and total type Ⅰ procollagen N-terminal peptide (P1NP) by electrochemiluminescence assay. Methods According to the Clinical and Laboratory Standards Institute (CLSI) "CA8-A" document the appropriately healthy people, who were divided into three groups (men, premenopausal women, and postmenopausal women) by sex and pre- or postmenopausal status were screened. The levels of fasting serum of OCN,β-CTx, tPINP were detected by Roche Modular E170 electrochemical immunoassay. Results 393 appropriately healthy people consists of 112 men between the ages of 29 and 69 years, 148 premenopausal women between the ages of 29 and 69 years, 133 postmenopausal women between the ages of 29 and 69 years. The levels of serum OCN, β-CTx, tP1NP in men group were (15.33±4.76) μg/L, (413±189) ng/L, (42.15±17.14) μg/L, respectively. The levels of serum OCN, β-CTx, tP1NP in premenopausal women group were (12.99±4.53) μg/L, 265(30-820) ng/L, (36.43±14.23) μg/L, respectively. The levels of serum OCN, β-CTx, tP1NP in postmenopausal women group were (18.96±5.15) μg/L, (513±195) ng/L, 51.40 (8.98 -118.6)μg/L, respectively. Logarithmic transformation produced normal distributions for all markers but serum β-CTx of premenopausal women group and serum tPINP of postmenopausal women group. The 95% of the distribution intervals for serum OCN, β-CTx, tP1NP in men group was 6.00-24.66 μg/L, 43-783 ng/L, 9.06-76.24 μg/L, respectively. The 95% of the distribution intervals for serum OCN, β-CTx, tP1NP in premenopausal women group was 4.11-21.87 μg/L, 68-680 ng/L, 8.53-64.32 μg/L respectively. The 95% of the distribution intervals for serum OCN, β-CTx, tPl NP in postmenopausal women group were 8.87-29.05 βg/L, 131-900 ng/L, 21.32-112. 80 μg/L, respectively. Conclusions Compared with the reference intervals provided by manufacture, the reference intervals of three serum bone turnover markers established by our laboratory have great difference. Laboratory should pay attention to the reference intervals was cited.