Millner P A, Hays H C W, Vakurov A, Pchelintsev N A, Billah M M, Rodgers M A
IMSB/FBS, Garstang Building, University of Leeds, Leeds LS2 9JT, UK.
Semin Cell Dev Biol. 2009 Feb;20(1):34-40. doi: 10.1016/j.semcdb.2009.01.011. Epub 2009 Feb 4.
For fabrication of effective electrochemical biosensors, interfacing the biomolecular receptor with the underlying transducer represents a critical step. The actual approach taken depends on the tethering layer covering the transducer, which is typically either a conducting polymeric matrix, or a thin film, such as an alkanethiol monolayer. Non-specific immobilisation methods can be either covalent, or non-covalent affinity attachment, with multipoint electrostatic attachment of the sensing biomolecule to either a polyanionic or polycationic layer representing the most common approach. Many specific affinity immobilisation strategies exist, but the majority make use of one of two binding systems. The first relies on the specific and strong affinity between biotin and proteins of the avidin family, with both bioreceptor and transducer bearing pendant biotins and avidin used as the crosslinker. The second approach employs a metal chelating group on the transducer to which can be bound a polyhistidine tag present on the N- or C-terminus of the receptor protein and which can be introduced genetically, when the expression sequence for a recombinant proteins is designed.
对于有效的电化学生物传感器的制造而言，将生物分子受体与底层换能器连接起来是关键步骤。所采用的实际方法取决于覆盖换能器的连接层，该连接层通常要么是导电聚合物基质，要么是薄膜，如烷硫醇单分子层。非特异性固定方法可以是共价的，也可以是非共价亲和附着，传感生物分子与聚阴离子或聚阳离子层的多点静电附着是最常见的方法。存在许多特异性亲和固定策略，但大多数利用两种结合系统之一。第一种依赖于生物素与抗生物素蛋白家族蛋白质之间的特异性强亲和力，生物受体和换能器都带有侧链生物素，抗生物素蛋白用作交联剂。第二种方法在换能器上使用金属螯合基团，受体蛋白的N端或C端存在的多组氨酸标签可以与之结合，当设计重组蛋白的表达序列时，可以通过基因方式引入该标签。
