Stoltenburg R, Reinemann C, Strehlitz B
Department of Environmental Biotechnology, Centre for Environmental Research Leipzig-Halle GmbH (UFZ), Permoserstr. 15, 04318 Leipzig, Germany.
Anal Bioanal Chem. 2005 Sep;383(1):83-91. doi: 10.1007/s00216-005-3388-9. Epub 2005 Oct 19.
Aptamers are ssDNA or RNA oligonucleotides with very high affinity for their target. They bind to the target with high selectivity and specificity because of their specific three-dimensional shape. They are developed by the so-called Systematic Evolution of Ligands by Exponential Enrichment (SELEX) process. We have modified this method in two steps-use of fluorescent labels for DNA quantification and use of magnetic beads for target immobilization. Thus, radioactive labelling is avoided. Immobilization on magnetic beads enables easy handling, use of very small amounts of target for the aptamer selection, rapid and efficient separation of bound and unbound molecules, and stringent washing steps. We have called this modified SELEX technology FluMag-SELEX. With FluMag-SELEX we have provided a methodological background for our objective of being able to select DNA aptamers for targets with very different properties and size. These aptamers will be applied as new biosensor receptors. In this work selection of streptavidin-specific aptamers by FluMag-SELEX is described. The streptavidin-specific aptamers will be used to check the surface occupancy of streptavidin-coated magnetic beads with biotinylated molecules after immobilization procedures.
适体是对其靶标具有非常高亲和力的单链DNA或RNA寡核苷酸。由于其特定的三维形状,它们以高选择性和特异性与靶标结合。它们是通过所谓的指数富集配体系统进化(SELEX)过程开发的。我们分两步对该方法进行了改进——使用荧光标记进行DNA定量以及使用磁珠固定靶标。因此,避免了放射性标记。固定在磁珠上便于操作,在适体筛选中可使用极少量的靶标,能快速有效地分离结合和未结合的分子,并且有严格的洗涤步骤。我们将这种改进的SELEX技术称为FluMag-SELEX。通过FluMag-SELEX,我们为能够针对具有非常不同性质和大小的靶标选择DNA适体这一目标提供了方法学背景。这些适体将用作新型生物传感器受体。在这项工作中,描述了通过FluMag-SELEX选择链霉亲和素特异性适体的过程。链霉亲和素特异性适体将用于检查固定程序后用生物素化分子包被的链霉亲和素磁珠的表面占有率。