Antibody characteristics for a continuous response fiber optic immunosensor for theophylline.
A self-contained fiber optic immunosensor was developed to measure continuously theophylline concentrations. The analytical signal was derived from the non-radiative energy transfer quench of fluorescence following binding of a fluorescence donor, B-phycoerythrin labeled theophylline, to an energy acceptor, Texas Red labeled antibody. Increases in free theophylline analyte concentrations resulted in a shift in antibody binding equilibrium between labeled and unlabeled theophylline that elicited a proportional increase in the fluorescence. The selection criteria for a monoclonal antibody as a molecular recognition element, and the optimization of labeling conditions to maximize the dynamic range and minimize sensor response time are described. Under one or more Texas Red labeling conditions, five antibody clones exhibited significant quenching when mixed with labeled analyte and also demonstrated 95% or greater reversible binding to labeled analyte. Two clones failed to exhibit fluorescence quenching when mixed with labeled analyte. The response time of the indicator chemistry system was dependent on the dissociation rate constant of the antibody. The equilibrium response time of intact sensors was limited by analyte diffusion across the containment membrane.
Hanbury
C M
CM
Department of Pathology and Biochemistry, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298-0597, USA.
Miller
W G
WG
Harris
R B
RB
D000937:Antigen-Antibody Reactions; D015374:Biosensing Techniques; D005336:Fiber Optic Technology; D005456:Fluorescent Dyes; D007700:Kinetics; D055100:Optical Fibers; D013050:Spectrometry, Fluorescence; D013806:Theophylline / Q000031:analogs & derivatives / Q000032:analysis*; D014966:Xanthenes
D005456:Fluorescent Dyes; D014966:Xanthenes; C034657:Texas red; D013806:Theophylline