Clapp Aaron R, Goldman Ellen R, Mattoussi Hedi
US Naval Research Laboratory, Optical Sciences Division, Code 5611, 4555 Overlook Avenue SW, Washington, DC 20375, USA.
Nat Protoc. 2006;1(3):1258-66. doi: 10.1038/nprot.2006.184.
We provide a detailed protocol for designing water-soluble CdSe-ZnS quantum dots (QDs) based on cap exchange of the native hydrophobic shell with dihydrolipoic acid (DHLA) ligands, and the preparation of functional QD bioconjugates for use in immunoassays. Our conjugation strategy is based on non-covalent self-assembly between DHLA-capped QDs and protein appended with either an electrostatic attachment domain (namely, the basic leucine zipper) or a polyhistidine tag. These bioconjugates combine the properties of the QD and attached biomolecule to create structures with desirable luminescent and biologically specific properties. This method also allows the preparation of mixed surface conjugates, which results in the conjugates gaining multiple biological activities. Conjugation of DHLA-capped QDs to maltose binding protein (MBP), the immunoglobulin-G-binding beta2 domain of streptococcal protein G (PG) and avidin will be described. MBP and PG were modified by genetic fusion with either a charged leucine zipper or a polyhistidine interaction domain.
我们提供了一份详细的实验方案，用于基于用二氢硫辛酸（DHLA）配体对天然疏水壳进行帽交换来设计水溶性CdSe-ZnS量子点（QD），以及制备用于免疫测定的功能性QD生物偶联物。我们的偶联策略基于DHLA封端的量子点与附加有静电附着域（即碱性亮氨酸拉链）或多组氨酸标签的蛋白质之间的非共价自组装。这些生物偶联物结合了量子点和附着生物分子的特性，以创建具有理想发光和生物学特异性特性的结构。该方法还允许制备混合表面偶联物，这导致偶联物获得多种生物活性。将描述DHLA封端的量子点与麦芽糖结合蛋白（MBP）、链球菌蛋白G（PG）的免疫球蛋白G结合β2结构域和抗生物素蛋白的偶联。MBP和PG通过与带电荷的亮氨酸拉链或多组氨酸相互作用域进行基因融合来修饰。
