A. V. El’skaya

D1 protein – an effective substitute for immunoglobulins in ELISA for the detection of photosynthesis inhibiting herbicides

Abstract A sensitive and inexpensive method of photosynthesis inhibiting herbicide detection, based on modification of the well-known enzyme-linked immunosorbent assay (ELISA) is described. The herbicide binding protein of chloroplasts (D1 protein) is used in ELISA as a substitute for specific immunoglobulins. Among the advantages of this innovation are simplicity of D1 protein preparation, its high stability, and group specificity for the photosynthesis inhibiting herbicides. Combination of the method proposed with the herbicide preconcentration on solid-phase cartridges allows to achieve the detection limit required by EC directives for drinking water quality. Several sorbents for solid-phase extraction (SPE) can be used for the selective analysis of different types of the photosynthesis inhibiting herbicides.

An amperometric glutamate biosensor for monitoring glutamate release from brain nerve terminals and in blood plasma

An excess of the excitatory neurotransmitter, glutamate, in the synaptic cleft during hypoxia/ischemia provokes development of neurotoxicity and originates from the reversal of Na+-dependent glutamate transporters located in the plasma membrane of presynaptic brain nerve terminals. Here, we have optimized an electrochemical glutamate biosensor using glutamate oxidase and developed a biosensor-based methodological approach for analysis of rates of tonic, exocytotic and transporter-mediated glutamate release from isolated rat brain nerve terminals (synaptosomes). Changes in the extracellular glutamate concentrations from 11.5 ± 0.9 to 11.7 ± 0.9 μΜ for 6 min reflected a low tonic release of endogenous glutamate from nerve terminals. Depolarization-induced exocytotic release of endogenous glutamate was equal to 7.5 ± 1.0 μΜ and transporter reversal was 8.0 ± 1.0 μΜ for 6 min. The biosensor data correlated well with the results obtained using radiolabelled L-[14C]glutamate, spectrofluorimetric glutamate dehydrogenase and amino acid analyzer assays. The blood plasma glutamate concentration was also tested, and reliability of the biosensor measurements was confirmed by glutamate dehydrogenase assay. Therefore, the biosensor-based approach for accurate monitoring rates of tonic, exocytotic and transporter-mediated release of glutamate in nerve terminals was developed and its adequacy was confirmed by independent analytical methods. The biosensor measurements provided precise data on changes in the concentrations of endogenous glutamate in nerve terminals in response to stimulation. We consider that the glutamate biosensor-based approach can be applied in clinics for neuromonitoring glutamate-related parameters in brain samples, liquids and blood plasma in stroke, brain trauma, therapeutic hypothermia treatment, etc., and also in laboratory work to record glutamate release and uptake kinetics in nerve terminals.

Selective Determination of Heavy Metal Ions with Sensors Coupled to Immobilised Enzymes

Selective biosensor systems for determination of heavy metal ions based on ion sensitive field effect transistors and thin-film interdigitated planar (conductometric) electrodes have been developed. Glucose oxidase, alcohol oxidase and butyril cholinesterase immobilised on the transducer surfaces have been used as bioactive elements for preparation of Ag+, Hg2+ and Pb2+ sensitive biosensors. The biosensors developed exhibit a dynamic range of 1 – 100 μM for these heavy metal ions. The performance characteristics of the biosensors are discussed.