A simple, sensitive and reproducible radioimmunoassay for determination of thymidine in biological fluids was developed. The technique is based on competition between 3H-thymidine and unlabelled thymidine for antithymidine antibodies. The cross-reactivity of structurally related compounds with the antibody was mostly negligible, except for deoxyuridine. The sensitivity of the assay was 10(-7) M. This radioimmunoassay may be used for measuring content of thymidine in blood serum and urine. A method for direct determination of thymidine in urine was not described before.