Plüddemann Annette, Neyen Claudine, Gordon Siamon, Peiser Leanne
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford, OX1 3RE, UK.
J Immunol Methods. 2008 Jan 1;329(1-2):167-75. doi: 10.1016/j.jim.2007.10.002. Epub 2007 Oct 25.
In order for macrophages to perform their numerous homeostatic, immunological and tissue remodeling functions they are required to express a broad repertoire of cell-surface receptors. These receptors are particularly important for their host-defense functions in the recognition of foreign pathogens. Delineation of the particular functions of specific receptors requires the identification of ligands recognized by the receptor. We have developed a sensitive, high throughput, solid-phase assay for the detection of ligands for the class A macrophage scavenger receptor (SR-A). Post-nuclear cell lysate from murine bone marrow-derived macrophages is used as a source of receptor and specific ligand binding to SR-A is detected with a monoclonal antibody for SR-A. This assay has been used effectively to identify protein ligands for SR-A on the surface of the bacterium Neisseria meningitidis (Peiser, L. et al. [Peiser, L., Makepeace, K., Pluddemann, A., Savino, S., Wright, J.C., Pizza, M., Rappuoli, R., Moxon, E.R., Gordon, S., 2006. Identification of Neisseria meningitidis nonlipopolysaccharide ligands for class A macrophage scavenger receptor by using a novel assay. Infect. Immun. 74, 5191-5199]). In this paper we describe the method in detail and define the specific variables governing the assay.
为了使巨噬细胞发挥其众多的稳态、免疫和组织重塑功能，它们需要表达多种细胞表面受体。这些受体对于它们在识别外来病原体时的宿主防御功能尤为重要。确定特定受体的特定功能需要识别该受体所识别的配体。我们开发了一种灵敏、高通量的固相检测方法，用于检测A类巨噬细胞清道夫受体（SR-A）的配体。来自小鼠骨髓来源巨噬细胞的核后细胞裂解物用作受体来源，并用抗SR-A单克隆抗体检测与SR-A特异性结合的配体。该检测方法已有效地用于鉴定脑膜炎奈瑟菌表面SR-A的蛋白质配体（Peiser, L.等人[Peiser, L., Makepeace, K., Pluddemann, A., Savino, S., Wright, J.C., Pizza, M., Rappuoli, R., Moxon, E.R., Gordon, S., 2006. 通过一种新型检测方法鉴定A类巨噬细胞清道夫受体的脑膜炎奈瑟菌非脂多糖配体。感染与免疫。74, 5191-5199]）。在本文中，我们详细描述了该方法，并确定了控制该检测的特定变量。
