This paper describes the development of a fiber-optic chemical sensor based on the principle of competitive-binding fluorescence immunoassay. Rabbit immunoglobln G (IgG) is covalently Immobilized on the distal sensing tip of a quartz optical fiber. The sensor is exposed to fluorescein Isothiocyanate (FITC) labeled and unlabeled anti-rabbit IgG. The 488-nm line of an argon-ion laser provides excitation of sensor-bound analyte. This results in fluorescence emission at the optical fiber‧s sensing tip. Sensor response is inversely proportional to the amount of unlabeled anti-IgG In the sample. Limits of detection (LOD) vary with Incubation time, sample size, and measurement conditions. For 10-μL samples, typical LOD are 25 fmol of unlabeled antibody In a 20-min Incubation period. These results Indicate that each fiber-optic fluoroimmunosensor can be constructed to perform a single sensitive, rapid, low-volume immunoassay, in in situ or benchtop applications. © 1987, American Chemical Society. All rights reserved.

This paper describes the development of an antibody-based fiber-optic sensor designed to detect benzo[a]pyrene (BP) metabolites, in this case r-7,t-8,9,c-10-tetrahydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene (benzopyrenetetraol or BPT), a naturally fluorescent hydrolysis product of DNA adducts that forms during BP metabolism. The sensor's principle of operation is based on diffusion of BPT Into the fiber's reagent phase where it complexes with monoclonal antl-BPT antibody. Reagent phase is immobilized Inside a reusable sensing tip. It consists of either membrane-entrapped liquid antibody or antibody Immobilized on 7-μm silica beads. The antibody binds and concentrates BPT in the fiber's viewing region until equilibrium with the sample is achieved. Typical measurements are performed in 15 min under nonequilibrium conditions. The 325-nm line of a He:Cd laser provides excitation of the immune complex. The Intensity of the resulting fluorescence emission is directly proportional to the amount of BPT in the sample. Sensor response rate varies with BPT concentration in the sample and, under certain circumstances, with antibody concentration in the sensor. At high antibody concentrations, sensor response rate is linear for about 2 h. For 15-min incubations, typical limits of detection are 5 X 10-10M. Significant chemical selectivity is also demonstrated despite the presence of structurally and spectrally similar polynuclear aromatic (PNA) interferents. These results indicate that the fluorolmmunosensor (FIS) is capable of performing highly sensitive and selective immunoassays and could be useful as a screening instrument for BP metabolites in biological samples. Furthermore, the Information provided by this work should be generally applicable to the development of devices for chemically similar systems. © 1988, American Chemical Society. All rights reserved.

The speed and specificity characteristic of immunochemical complex formation has encouraged the development of numerous antibody-based analytical techniques. The scope and versatility of these established methods can be enhanced by combining the principles of conventional immunoassay with laser-based fiber-optic fluorimetry. This merger of spectroscopy and immunochemistry provides the framework for the construction of highly sensitive and selective fiber-optic devices (fluoroimmuno-sensors) capable of in-situ detection of drugs, toxins, and naturally occurring biochemicals. Fluoroimmuno-sensors (FIS) employ an immobilized reagent phase at the sampling terminus of a single quartz optical fiber. Laser excitation of antibody-bound analyte produces a fluorescence signal which is either directly proportional (as in the case of natural fluorophor and ‘antibody sandwich’ assays) or inversely proportional (as in the case of competitive-binding assays) to analyte concentration. Factors which influence analysis time, precision, linearity, and detection limits include the nature (solid or liquid) and amount of the reagent phase, the method of analyte delivery (passive diffusion, convection, etc.), and whether equilibrium or non-equilibrium assays are performed. Data will be presented for optical fibers whose sensing termini utilize: (1) covalently-bound solid antibody reagent phases, and (2) membrane-entrapped liquid antibody reagents. Assays for large-molecular weight proteins (antigens) and small-molecular weight, carcinogenic, polynuclear aromatics (haptens) will be considered. In this manner, the influence of a system's chemical characteristics and measurement requirements on sensor design, and the consequence of various sensor designs on analytical performance will be illustrated. © 1988 SPIE.