Buckingham Steven David, Pym Luanda, Sattelle David Barry
Department of Human Anatomy and Genetics, University of Oxford, UK.
Methods Mol Biol. 2006;322:331-45. doi: 10.1007/978-1-59745-000-3_23.
The Xenopus laevis oocyte offers one of the most convenient expression systems for assaying the actions of candidate ligands on cloned ionotropic neurotransmitter receptors (also known as ligand-gated ion channels [LGICs]). Their large size makes injection of complementary ribonucleic acid or complementary deoxyribonucleic acid and electrophysiological recording very easy. Furthermore, Xenopus oocytes translate messages very efficiently, resulting in the detection of large-amplitude ligand-induced currents from expressed, recombinant LGICs. Compared to other electrophysiological techniques, recording from oocytes is not difficult and requires only a basic electrophysiological recording setup. Oocytes can be used for two-electrode voltage clamp, as well as cell-attached patch and inside- or outside-out patch clamp recordings. A variety of protocols allows the experimenter to determine the actions of ligands on cloned receptors and parameters, such as their affinity, efficacy, rates of association and desensitization, and reversibility, to be estimated. Here, we present protocols for using Xenopus oocytes in assaying candidate ligands acting against cloned targets of drugs and pesticides.
非洲爪蟾卵母细胞为检测候选配体对克隆的离子型神经递质受体（也称为配体门控离子通道[LGICs]）的作用提供了最便捷的表达系统之一。它们的体积大使得注射互补核糖核酸或互补脱氧核糖核酸以及进行电生理记录非常容易。此外，非洲爪蟾卵母细胞能非常高效地翻译信息，从而能够检测到来自表达的重组LGICs的大幅度配体诱导电流。与其他电生理技术相比，从卵母细胞进行记录并不困难，只需要一个基本的电生理记录装置。卵母细胞可用于双电极电压钳，以及细胞贴附式膜片钳和内面向外或外面向内的膜片钳记录。多种实验方案使实验者能够确定配体对克隆受体的作用，并估计其亲和力、效力、结合和解敏速率以及可逆性等参数。在这里，我们介绍使用非洲爪蟾卵母细胞检测作用于药物和农药克隆靶点的候选配体的实验方案。
