A highly sensitive enzyme immunoassay for evaluation of 2'-deoxycytidine plasma level as a prognostic marker for breast cancer chemotherapy.

A highly sensitive competitive enzyme immunoassay (EIA) has been developed and validated for the determination of the plasma level of 2'-deoxycytidine (dCyd), the potential prognostic marker for breast cancer chemotherapy. This assay employed a monoclonal antibody that recognizes dCyd with a high specificity, and 5'-succinyl-dCyd (5'sdCyd) conjugate of bovine serum albumin (5'sdCyd-BSA) immobilized onto microplate wells as a solid phase. The assay involved a competitive binding reaction between dCyd, in plasma sample, and the immobilized 5'sdCyd-BSA for the binding sites of the anti-dCyd antibody. The bound antibody was quantified with horseradish peroxidase-labeled anti-immunoglobulin second antibody and 3,3',5,5'-tetramethylbenzidine as a peroxidase substrate. The concentration of dCyd in the sample was quantified by its ability to inhibit the binding of the antibody to the immobilized 5'sdCyd-BSA and subsequently the color formation in the assay. The assay limit of detection was 8 nM and the effective working range at relative standard deviations (R.S.D.s) of