A novel synthetic peptide spacer designed on a gold film is introduced for use in surface plasmon resonance (SPR) sensing. The peptide was a specially designed sequence of amino acids, synthesized by the Fmoc-solid-phase chemistry. The peptide was adsorbed on the gold film from an aqueous solution via its four thiol groups, forming a self-assembled negatively charged monolayer. The monolayer contained carboxyl groups, which were activated by the EDC/NHS technique. It was successfully used as a matrix (2 - 4 nm thick) for covalent immobilization of fusion protein, which included C-terminal fragment of human 5-hydroxytryptamine transporter (molecular weight approximately 21 kDa). The reaction between the immobilized protein and antibodies was monitored by SPR means. The matrix did not cause degradation of immobilized components and steric hindrances to mass transport, and also demonstrated low nonspecific binding to antibodies. Besides, the matrix could be regenerated without decreasing SPR response to the reaction. Along with the ability to immobilize high weight molecules, which are unable to enter a conventional CM-dextran matrix due to steric hindrances to mass transport, the peptide matrix has a number of advantages over the CM-dextran matrix, namely, simplicity in preparation, low cost, and much shorter time needed for preparation. The peptide spacer matrix can be widely used not only in SPR, but also in other analytical techniques that require immobilization of proteins on metal surfaces, such as interferometry, piezoelectric detection, scanning tunneling microscopy.
An optical biosensor has been developed for detection of pesticides, based on surface plasmon resonance (SPR) technique. Concentration of the pesticides was measured in liquid or gas. We specially originated organic film on a disposable element. A setup on the base of Kretschmann arrangement was improved by using a computer-controlled angular scanning system. The detection concentration limit of dinitrophenole (DNP) was 10-9 M. Some samples exhibited effect down to 10-11 M of DNP. The results obtained provide reason for further development of SPR sensor as applied to pesticides monitoring.
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