Development of a fluoroimmunosensor for theophylline using immobilised antibody

A flowthrough theophylline fluoroimmunosensor with an antibody covalently i mmobilised on a solid support has been developed. The immobilisation techni que proposed in this paper used protein-A on control pore glass (Protein A- CPG) in an immunoreactor and dimethylsuberimidate as a cross-linking agent. Several supports and cross-linking reagents were tested in order to obtain oriented immobilisation and thus efficiency of the immunological reaction and reusability of the immunosensor. The immunosensor performance character istics were established. The precision expressed as RSD, was 1.6%; the dete ction limit was 3 mu g l(-1); the immunoreactor lifetime was established in 80 assays and there were no interferences with structurally similar compou nds such as aminophylline, dihydroxypropyltheophylline and caffeine in the determination of the analyte. This fluoroimmunosensor was applied to determ ine theophylline in human serum samples from patients of the Puerta de Hier ro Hospital in Madrid. The results obtained show that there are no signific ant differences between the proposed immunosensor and the high-pressure liq uid chromatographic method with UV detection used by the Hospital, thus dem onstrating the validity of the method. (C) 2000 Elsevier Science B.V. All r ights reserved.