BACKGROUND:Biomarkers can aid in detecting and preventing clinical disease through the recognition of change in biological samples. The objective of this case-control study was to further the knowledge on the use of big toenail and hair samples as biomarkers for fluoride exposure. METHODS:A total of 48 participants from an endemic (IC) and a non-endemic (SC) fluorosis region were included. Big toenail and hair samples were collected from each participant and the fluoride concentration was determined. The results of 42 participants were reported: 21 participants (11 males and 10 females, mean age 15.66 + 2.61 years) from IC and 21 participants (11 males and 10 females, mean age 15.06 + 0.79 years) from SC. RESULTS:The mean fluoride concentration of big toenail (2.34 ± 0.26 mg/kg) and hair (0.24 ± 0.04 mg/kg) in the endemic region was significantly higher than the mean fluoride concentration of big toenail (0.98 ± 0.08 mg/kg) and hair (0.14 ± 0.02 mg/kg) in the non-endemic region (p < 0.001 and p = 0.004, respectively). The Receiver Operating Characteristic (ROC) analysis showed that the Area Under the Curve (AUC) value was 0.889 for big toenail (p < 0.001) and 0.762 for hair (p = 0.004) samples. The fluoride assay for big toenails exhibits greater observed accuracy than does the fluoride assay for hair. CONCLUSION:Nail and hair samples can serve as biomarkers to detect biological fluoride exposure according to the data of this pilot study. Nevertheless, hair is less sensitive and specific as a biomarker when AUC values of big toenail and hair samples were compared.