By Harlow Shapley
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22 Biosensors based on self-assembled monolayers A higher sensitivity can be achieved by using ssDNA label probes, which are tagged with an electroactive species. The use of a "sandwich" assay format is specially advantageous in this context because it obviates the need to tag the oligonucleotide target probe, it does not require a washing step, thus giving rise to a "real time" response and it can be directly coupled to a polymerase chain amplification reaction . This method has, however, a serious drawback because of the extremely small a m o u n t of label oligonucleotide probes captured on the electrode surface, which gives rise to an accordingly small electrochemical response.
In this case, surface-bound reactive groups not only provide anchoring sites for protein attachment, but also build a "carpet" of surface charges, which can promote a direct electron transfer reaction between surface-bound proteins, such as cytochrome-c  and cytochrome-c oxidase  and the electrode surface. D. Schlereth mediator) is required to wire the protein to the electrode surface, although their use is not desirable for biosensors used as analytical tools in food technology or for clinical purposes.
1 Multipoint a t t a c h m e n t o f p r o t e i n s to gold e l e c t r o d e surfaces modified w i t h an alkylthiol SAM Most redox proteins, either in solution or adsorbed at an electrode surface, show a poorly defined electrochemical response at electrode surfaces because their redox site is deeply buried within their polypeptide backbone, which acts as an electrical insulator hindering the electron transfer reaction . However, when the redox site is close to the protein surface and the protein approaches (or is immobilized onto) the electrode surface, such that the redox site faces the surface, the rate of the electron transfer reaction is sufficiently high to give rise to a direct (non-mediated) quasi-reversible electrochemical response .