Objective. Monitoring inflammatory diseases and osteoclastogenesis with osteopontin (OPN), osteoprotegerin (OPG), total soluble receptor activator of nuclear factor kappa B ligand (total sRANKL) and high-sensitivity C-reactive protein (hsCRP) has recently attracted increased interest. The purpose of our study was to determine reference intervals, variability caused by sampling time, biological variation and stability after repeated freeze-thaw cycles of circulating levels of OPN, OPG, total sRANKL and hsCRP. Material and methods. Plasma OPN and plasma OPG concentrations were determined with sandwich ELISA; serum total sRANKL concentration was determined using a two-site sandwich ELISA; and hsCRP was analysed by turbidimetry in 300 Danish blood donors (183 M and 117 F) with a median age of 43 years (range 18-64 years). Variability due to biological variation and sampling time was studied in serial samples from 38 healthy subjects. Results. The 95th percentiles in the donors were 76 µg/L for OPN, 4.2 pmol/L for OPG, 40.2 nmol/L for total sRANKL and 12 mg/L for hsCRP. The overall medians for both genders were 51 µg/L, 2.2 pmol/L, 0.66 nmol/L and 1.0 mg/L, respectively. We found a significant correlation between hsCRP and OPN (rho = 0.173; p<0.003). The biological within-subject variations were calculated to be 8.2 % for OPN, 8.8 % for total sRANKL and 50 % for hsCRP. Conclusions. Reference intervals have been established with a high analytic performance for OPN and an acceptable analytic performance for OPG and total sRANKL. The study revealed low biological variation for OPN and total sRANKL and high biological variation for hsCRP.