Nickel is widely used in many industries. It is known as an immunotoxic, neurotoxic, pulmonary toxic, nephrotoxic, hepatotoxic, and carcinogenic agent. Biological monitoring is a useful tool to assess the internal exposure to toxic compounds. Because of trace concentration of analytes in biological samples and matrix interferences, the preparation of such complicated samples is necessary prior to analysis. This study aimed to optimize the parameters affecting solid phase extraction (SPE) procedure and compare the enrichment recoveries of sorbents XAD-7 and chromosorb 105 for preconcentration and separation of nickel in urine, hair and nail samples. Solid phase extraction using both resins was optimized regarding the sample pH, ligand concentration, loading flow rate, elution solvent, sample volume, elution volume, amount of resins, and matrix interferences. The optimized procedure was validated based on the "within-day" and "day-to-day" reproducibility experiments. The proposed method was applied for pre-treatment of urine, hair and nail samples. Nickel was extracted with recoveries in the range of 94-100%. A good reproducibility and repeatability was obtained for optimized method using both resins under the existing conditions. The concentration of nickel in workers’ biological samples was determined using two sorbents. There was no significant difference between obtained recoveries for XAD-7 and chromosorb 105. This optimized method can be successful in simplifying sample preparation for trace residue analysis of Ni in different matrices for evaluation of occupational and environmental exposures. Resin XAD-7 is recommended for next experiments because of its lower cost and simpler preconditioning compared to chromosorb 105.