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We describe a novel pyruvate selective sensor in which corn tissue, as the biocatalytic layer, is coupled to a fluorescent indicator-based fiber-optic CO2 optrode. This sensor showed shorter response times and a lower detection limit without loss of enzymatic stability as compared to a corn tissue-based pyruvate selective membrane electrode previously studied.

A dual-working electrode (K1 and K2) multienzyme sensor, made of carbon paste, glucose oxidase (GOD) membrane (on K1) and invertase (INV)/mutarotase (MUT)/GOD membrane (on K2), is proposed for simultaneous determination of glucose and sucrose. Interference to the K2 caused by H2O2 produced in the enzyme reaction on K1 can be eliminated by depositing a catalase (CAT) layer between K1 and K2. Acceleration of glucose mutarotation by MUT was found to be three times faster than by phosphate buffer. Glucose was determined directly from the response of K1, while sucrose was computed based on a formula derived from a two-point calibration method. The maximum concentration of sucrose that could be determined depended strongly on the concentration of coexisting glucose. The activities of immobilized GOD, INV, MUT, and CAT were examined individually and found stable at room temperature for at least 1 month. The final loss of sucrose response activity was found to be due to denaturation of INV.

The preparation and characterization of rabbit and bovine IgG-coated membranes is discussed. The performance of these membranes in a competitive binding assay for rabbit IgG and in a sandwich assay for rabbit anti-bovine IgG were used to judge the potential usefulness of these membranes as solid phases for enzyme immunosensors. In particular, these membranes were intended for use in enzyme immunosensors based on the ammonia gas-sensing electrode which employed urease labeled immunoreagents.

An investigation of the antibody sensing properties of potentiometric polymer membrane electrodes incorporating antigens shows that at least two categories of such electrodes are possible. When the immobilized antigen has no ionophoric character of its own, antibody response can be obtained only upon conjugation of the antigen to an ionophore. The second category, corresponding to a new class of electrodes, yields potentiometric antibody response in the absence of a conjugated ionphore. These effects are illustrated with electrodes for antidigoxin, anti-DNP, and antiquinidine, respectively.

A mixture of intact bacterial cells and an enzyme extract is used in conjunction with an ammonia gas sensor to construct a new type of hybrid membrane electrode. The hybrid electrode measures the coenzyme NAD+ by means of a biocatalytic sequence that produces ammonia in stoichiometric amounts. Calibration plots are linear from approximately 2.5 x 10-4 to 2.5 x 10-3M NAD+ with a slope of -49 mv/decade. The electrode has a useful lifetime of at least one week. The present work demonstrates that hybrid electrodes can be developed for the measurement of substances which require multi-step sequences to form final electroactive products and, thus, extends the range of applicability of bioselective membrane electrodes.

Results are presented for potentiometric ammonia gas-sensors which show that the response times increase significantly with age of the sensor when ammonium picrate is used as internal electrolyte and the samples have greater osmolarities than the internal electrolyte solution. A build-up of ammonium picrate crystals on the inner side of the gas-permeable membrane is responsible for this increasein response time. The use of ammonium chloride as internal electrolyte provides constant response times indepent of sample osmotic strength, and is recommended as a means of ensuring that optimal dynamic response characteristics are maintained.

A simple, sensitive and selective method is proposed for the determination of nicotinamide adenine dinucleotide phosphate (NADPH) in the concentration range 10−4–10−7 mol l−1 (90 μg—90 ng ml−1). The method is based on monitoring with a silver sulfide membrane electrode the concentration of the reduced glutathione released at pH 8 by reaction of oxidized glutathione with NADPH in the presence of glutathione reductase. Calibration data for 10−4–10−7 mol l−1 range of NADPH yielded a least-squares equation of (ΔE, mV) = (36.65 ± 0.15) log C + 264.93 ± 0.1 with a standard error of 0.4 mV. Other common nucleotides including NADP+ do not interfere.

The selectivity and response of neutral carrier based polymer membrane electrodes are investigated via exchange current measurements for systems containing valinomycin, dibenzo-18-crown-6, and plasticizer alone in a polyvinyl chloride matrix. Using a transient galvanostatic step method, apparent exchange current densities of 1.3 X 10−3 A/cm2, 5.4 X 10−6 A/cm2, and 2.2 X 10−9 A/cm2 were obtained with K+ as the primary ion for the three types of membranes, respectively. Preliminary results indicate that the exchange current data obtained with this technique are complementary to the potentiometric response observed for the membranes studied.

Antibody levels in the μg ml -1 range are selectively and rapidly determined in an automated flow system which employs an Ag 2 S membrane electrode sensor in conjunction with immobilized immunoabsorbent. Experiments with anti-benzoate antibody in the presence of excess γ-globulin or whole serum demonstrate the operating characteristics of the system in terms of selectivity, precision and dynamic response.

An ammonia gas-sensing membrane electrode is used as a potentiometric sensor for the assay of leucine aminopeptidase. The method utilizes hydrolysis of L-leucinamide to release ammonia which is detected by the electrode through initial rate measurements and related to the enzymatic activity. A within-run relative imprecision of ± 2.5% was established for the method with a working range up to 70 mU ml−1 enzyme. Recovery experiments in pooled serum showed an average recovery of 96.6% added enzyme. Good agreement with a spectrophotometric method was obtained for synthetic laboratory samples.

The coenzyme pyridoxal-5'-phosphate (PLP) is deiermined by in situ measurement of the rate of ammonia production using a potentiometric gas-sensing membrane electrode. It is shown that the initial rate of ammonia liberation from L-tryptophan by action of tryptophanase apoenzyme and the coenzyme can be linearly related to PLP levels in the nanogram range with minimal interference from related compounds. Calibration data for the 1 × 10 -7 –2.5 × 10 -6 M range for PLP yielded a least-squares equation of rate (mV min -1 ) = (0.64 ± 0.01) C — 0.01 ± 0.02 with a standard error of 0.02 mV min -1 , where concentration, C, is expressed in units of 10 -7 mol l -1 .

An iodide membrane electrode, in conjunction with rat liver microsomes, is used to measure thyroxine at sub-micromolar levels. The microsomes serve as biocatalytic reagents which selectively remove iodide from the hormone. The amount of iodide liberated, as measured by the electrode, is proportional to the initial thyroxine concentration. The electrode can further be employed to directly monitor the rate of deiodination of the hormone with advantages of speed and convenience over existing techniques.

A newly developed pyruvate selective membrane electrode using plant tissue as biocatalyst is described. Comparison studies of electrodes which employ corn kernel portions as biocatalyst with similar electrodes based on isolated enzyme show that the former achieve longer lifetimes and enhanced activities with no loss of selectivity. This improvement is attributed to stabilization of the biocatalytic activity by the plant tissue matrix.

A two-step enzyme sequence is coupled to a potentiometric gas sensor to prepare an electrode for D-gluconate, a substrate not measurable by conventional enzyme electrodes. The experimental variables and kinetic considerations for enzyme sequence electrodes are detailed using the gluconate system as a model. This research suggests that multi-step enzyme catalysis could usefully extend the range of membrane electrodes to new molecular species.

A glutamine-selective sensor consisting of porcine kidney tissue immobilized at an ammonia gas electrode is utilized. It yields good precision and accuracy over the clinically important range of glutamine concentrations (10-4–10-2 M).

I. INTRODUCTION AND SCOPE The ion-selective electrode is an electrochemical sensor characterized by a selective response to a particular ionic species. The wording “selective” rather than “specific” is considered since it is very rare that an electrode responds to a single ion exclusive of all others. The ion-selective electrode is further described as a membrane sensor which responds in a potentiometric manner. That is, the potential which develops across an ion-selective membrane separating two solutions is measured at virtually zero current. This is a rather unique and intriguing event which has founded an expanding analytical field which has come of age within the last 15 years.

A new class of fully reversible biosensors, utilizing a layer of monoclonal antibodies trapped at the sensing surface of an immobilized-antigen membrane, is shown to be suited for continuous monitoring of antigen levels. The model sensor described gives a potentiometric response to the hapten 2,4-dinitrophenol (DNP) at micromolar concentration levels through competitive interaction with monoclonal anti-DNP antibodies held within the sensor tip. The proposed design yields a reusable, reagentless antigen sensor with good analytical precision and an operating lifetime of at least 17 days.

Trypsin activity is monitored in a simple kinetic method with an ammonia gas sensor. The method is based upon the hydrolysis of α-benzoyl-arginine amide resulting in the production of ammonia. Day-to-day precision range of ±1.9% to 4.4% and a within-run precision range of ± 1.8% to 4.2% were measured for the working range of 0.5 to 20 μg mL−1 trypsin. An average of 97 ± 3.4% added trypsin was recovered from rat pancreatic juice in recovery experiments.

A novel potentiometric membrane electrode with selective response to L-glutamate over the 2 × 10 -4 –1.3 × 10 -2 M concentration range is constructed by immobilizing slices of yellow squash tissue at a CO 2 gas sensor. This system represents the first successful use of intact plant materials as biocatalysts in the construction of bioselective potentiometric membrane electrodes; as a result, the range of future possibilities may be greatly extended. The glutamate-sensing electrode combines excellent selectivity characteristics over some 25 possible interferences tested, good reproducibility, and a useful lifetime of 7 days in aqueous samples.

A membrane electrode with selective response for the antibodies of the antigenic determinant dinitrophenol (DNP) has been devised by incorporating DNP—ion carrier conjugates in a polyvinyl chloride membrane matrix. Details of the preparative and immobilization procedures are provided along with an evaluation of optimum solution conditions for electrode use. The response mechanism of the antibody electrode is postulated to involve a “selectivity shift” effect.

The sensitivity of potentiometric methotrexate assays can be enhanced by the presence of antimethotrexate antibody, which acts to potentiate the inhibitory effect of methotrexate on enzyme activity. This property is demonstrated in an enzyme-modified determination of methotrexate with a PCO2 membrane electrode in a cyclic scheme using the enzymes dihydrofolate reductase and 6-phosphogluconic dehydrogenase.

The use of intact bacterial cells or tissue slices of plant and animal origin as immobilized biocatalysts has extended the possible range of potentiometric bioselective membrane electrodes beyond that of conventional enzyme electrodes. The use of such materials as biocatalysts offers advantages in situations where isolated enzymes are not available or where multistep reaction paths are required. The resulting bioselective electrodes also offer exceptional ease of preparation, time stability, and low cost.

Direct antibody-sensing membrane electrodes have been developed by immobilizing ion-carrier immunogen conjugates in a liquid membrane matrix. The resulting potentiometric probes measure specific antibodies with high selectivity over nonspecific antibodies in the physiological pH range. The electrode response is shown to arise from the selective interaction of the antibody with the membrane-bound immunogen.

Publisher Summary All the tissue-based electrodes developed, whether from plant or animal sources, have utilized gas-sensing membrane sensors because of inherent advantages in simplicity of operation and excellent selectivity properties. Typically, a thin slice of tissue material is mechanically entrapped at the tip of the gas sensor, although fragile tissues may need to be stabilized with an immobilization procedure. This arrangement is no more tedious than the preparation of conventional enzyme electrodes and, often, is much simpler. This chapter provides an overview of the tissue electrodes. Mammalian tissues are a rich source of enzymes; indeed, many commercially available enzymes are isolated from such tissues. Thus, early development of tissue-based electrodes focused on the use of liver and kidney slices as biocatalysts, and, because of the ready availability of fresh organs from food animals in markets and slaughterhouses, initial experiments on such tissues employed those from bovine and porcine sources.

MEMBRANE electrodes based on living bacterial cells1 as biocatalysts have been shown to be useful sensors for amino acids and other biomolecules. We report here that we have now developed a highly effective membrane electrode for glutamine using slices of porcine kidney tissue as the biocatalytic layer. This tissue-based electrode yields excellent selectivity and sensitivity while having the additional advantages of simplicity and low cost. Further, the tissue-based electrode shows a much longer useful lifetime than comparable electrodes prepared with the isolated porcine enzyme.

The design and properties of a new enzyme electrode with response and selectivity for cyclic AMP(cAMP) are described. The cAMP sensor employs a double enzyme system held at the surface of an NH3-selective membrane electrode and has a detection limit of 5×10-9 moles cAMP as well as selectivity for cAMP over other nucleotides.

Ion-selective electrodes constructed with polyacrylamide(PAA) coupled to acyclic poly(oxyethylene) neutral carriers, tetraethylene glycol monododecyl ether (POE4) or hexaethylene glycol monododecyl ether (POE6), were reported. Nernstian responses were obtained to magnesium and barium ions in AN with the PAA–POE4 membrane and to copper(II) and silver(I) ions in DMSO with the PAA–POE6 membrane.

Des electrodes de Teflon comportant, aussi, un disque de platine, recouvert de polyacrylamide contenant un ionophore qui est un polyether macrocyclique, sont sensibles aux ions baryum, dans l'acetonitrile