Limit immunosensors

Immunosensors promise to become principal players ia chemical, diagnostic, and environmental analyses by the latter 1990s. Given the practical limits of immunosensors (low ppb or ng/mL to mid-pptr or pg/mL) and their portabiUty, the primary appHcation is expected to be as rapid screening devices ia noncentralized clinical laboratories, ia iatensive care faciUties, and as bedside monitors, ia physicians offices, and ia environmental and iadustrial settings (49—52). Industrial appHcations for immunosensors will also include use as the basis for automated on-line or flow-injection analysis systems to analyze and control pharmaceutical, food, and chemical processing lines (53). Immunosensors are not expected to replace laboratory-based immunoassays, but to open up new appHcations for immunoassay-based technology. 

The working conditions of the immunosensor (enzyme and antigen concentrations, dilutions of the antibodies, pH of the buffer solution) were found. The cholinesterase immobilized demonstrated the maximum catalytic activity in phosphate buffer solution with pH 8.0. The analytical chai acteristics of the sensor - the interval of the working concentrations and detection limit - have been obtained. The proposed approach of immunoassay made possible to detect 5T0 mg/ml of the bacterial antigen. 

Competitive immunoassays may also be used to determine small chemical substances [10, 11]. An electrochemical immunosensor based on a competitive immunoassay for the small molecule estradiol has recently been reported [11]. A schematic diagram of this immunoassay is depicted in Fig. 5.3. In this system, anti-mouse IgG was physisorbed onto the surface of an SPCE. This was used to bind monoclonal mouse anti-estradiol antibody. The antibody coated SPCE was then exposed to a standard solution of estradiol (E2), followed by a solution of AP-labeled estradiol (AP-E2). The E2 and AP-E2 competed for a limited number of antigen binding sites of the immobilized anti-estradiol antibody. Quantitative analysis was based on differential pulse voltammetry of 1-naphthol, which is produced from the enzymatic hydrolysis of the enzyme substrate 1-naphthyl phosphate by AP-E2. The analytical range of this sensor was between 25 and 500pg ml. 1 of E2. 

An immunosensor based on a competitive fluorescence energy-transfer immunoassay was reported by Anderson 105) for the measurement of phenytoin. Texas red-labeled antibody was incubated with a phenytoin derivative. On displacement of the derivative by the antigen, the change in the fluorescence signal was recorded. Detection limits approached 5 /iM with response times ranging from 5 to 30 min. 

A reversible, direct fluoroimmunosensor for human serum albumin (HS A) measurement has been described by Bright et al.(m> Antibody Fab fragments are first immobilized on small quartz plates by hinge-region thiols, and then dansylated. The immunosensor is formed by attaching the quartz plates with bound Fab to the distal end of a bifurcated fiber-optic probe, which transmits both the excitation and emission. Binding of ffSA to the immunosensor results in a three- to five-fold enhancement of dansyl fluorescence. The sensor can be reused up to 50 times, with a detection limit of about 1.8 x 10-8 M, and a somewhat limited dynamic range. 

Electrochemical immunosensors are a powerful tool for the analysis of antibacterials in food and different configurations have been presented during recent years. For example, an amperometric immunosensor was reported by Wu et al. [182], for penicillin quantification in milk, with a linear range from 0.25 to 3 ng/ml and a limit of detection of 0.3 pg/L [182]. Other types of transduction have been also explored, like a label-free impedimetric flow injection immunosensor for the detection of penicillin G. 

Electrochemical immunosensors are the most numerous of immunosensors. Their suitability for use in flow systems is limited by the two factors commented on above in dealing with fibre optic-based sensors, viz. the need for an incubation period and exceedingly long response times. 

Enantioselective, potentiometric membrane electrodes (EPMEs) are proposed for the potentiometric detection of the enantiomers [2,10]. The advantages of utilization of these electrodes over amperometric biosensors and immunosensors are a longer lifetime, a large working concentration range, no dilution required for the samples and possibility of decreasing of limit of detection by utilization of KC1 0.1 mol/L as internal solution [2],... 

Wherever an antibody assay is used now, there will be the prospect of producing an immunosensor for the same analyte. Therefore, the development of immunosensors is directed to obtain sensors with fast response, low detection limits, little preparation efforts, low price availability and high specificity. Thanks to the possibility of fabricating fast portable sensors, immunosensors now constitute a potential alternative to centralized and sophisticated bioanalytical systems. 

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