Membrane-bound and soluble forms of an NMDA receptor extracellular domain retain epitopes targeted in auto-immune encephalitis.
Membrane-bound and soluble forms of an NMDA receptor extracellular domain retain epitopes targeted in auto-immune encephalitis.
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DOI：
10.1186/s12896-018-0450-1
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发表时间：
2018-06-27
影响因子：
3.5
通讯作者：
Dessain SK
中科院分区：
文献类型：
--
作者：
Sharma R;Al-Saleem FH;Puligedda RD;Rattelle A;Lynch DR;Dessain SK
抗N-甲基-D-天冬氨酸受体脑炎（ANRE）是一种由抗N-甲基-D-天冬氨酸受体（NMDAR）自身抗体引起的潜在致死性疾病。如果在病程早期开始治疗，完全康复是可能的。检测脑脊液（CSF）中的ANRE抗体对诊断至关重要。ANRE相关IgG的测定通常依赖于用NMDAR基因瞬时转染的细胞。稳定表达致病性NMDAR表位的细胞系可以提高测定的标准化，并提供可用于商业固态测定系统的抗原。我们将GluN 1 NMDAR亚基（NR 1）的氨基末端结构域（ATD）表达为293 T细胞外质膜上的融合蛋白，产生了一个稳定的细胞群（293 T-ATD），该细胞群在流式细胞术和免疫荧光测定中被ANRE患者单克隆抗体识别。ATD融合蛋白还含有Myc标签和6XHIS标签，其提供用于免疫测定和抗原纯化的功能，以及TEV蛋白酶位点，其允许ATD结构域以基本上纯的形式从细胞特异性释放。从293 T ATD细胞系动员的ATD在ELISA结合测定中保持致病性ANRE表位。ANRE患者的CSF（3/4）和血清（4/4）也结合293 T-ATD细胞系，而正常CSF和血清不结合。293 T-ATD细胞系可能适用于多种形式以鉴定与ANRE相关的抗体，包括基于细胞的和可溶性抗原形式，并证明了产生用于研究、药物发现和临床诊断的复杂蛋白质的有用方法。本文的在线版本（10.1186/s12896-018-0450-1）包含补充材料，可供授权用户使用。
Anti-NMDA receptor encephalitis (ANRE) is a potentially lethal disease attributed to auto-antibodies against the N-methyl-D-aspartate receptor (NMDAR). Full recovery is possible if therapy is initiated early in the disease course. Detection of ANRE antibodies in the cerebrospinal fluid (CSF) is essential for diagnosis. The assays for ANRE-associated IgGs often rely on cells transiently transfected with NMDAR genes. A cell line that stably expresses pathogenic NMDAR epitopes could improve standardization of the assays and provide antigen that could be used in commercial solid state assay systems. We expressed the amino terminal domain (ATD) of the GluN1 NMDAR subunit (NR1) as a fusion protein on the outer plasma membrane of 293T cells, creating a stable cell population (293T-ATD) that is recognized by ANRE patient monoclonal antibodies in flow cytometry and immunofluorescence assays. The ATD fusion protein also contains a Myc tag and a 6XHIS tag, which provide functionality for immunoassays and antigen purification, and a TEV protease site, which allows the ATD domain to be specifically released from the cells in essentially pure form. ATD mobilized from the 293T ATD cell line maintained the pathogenic ANRE epitopes in ELISA binding assays. CSF (3/4) and sera (4/4) from ANRE patients also bound the 293T-ATD cell line, whereas normal CSF and sera did not. The 293T-ATD cell line is potentially adaptable to a variety of formats to identify antibodies associated with ANRE, including cell-based and soluble antigen formats, and demonstrates a useful method to produce complex proteins for research, drug discovery, and clinical diagnosis. The online version of this article (10.1186/s12896-018-0450-1) contains supplementary material, which is available to authorized users.
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DOI：
10.1016/j.jpeds.2012.10.011
发表时间：
2013-04
期刊：
The Journal of pediatrics
影响因子：
--
作者：
Armangue T;Titulaer MJ;Málaga I;Bataller L;Gabilondo I;Graus F;Dalmau J;Spanish Anti-N-methyl-D-Aspartate Receptor (NMDAR) Encephalitis Work Group
通讯作者：
Spanish Anti-N-methyl-D-Aspartate Receptor (NMDAR) Encephalitis Work Group
影响因子：
14.5
作者：
Kreye, Jakob;Wenke, Nina K.;Pruss, Harald
通讯作者：
Pruss, Harald
DOI：
10.1016/s1474-4422(15)00401-9
发表时间：
2016-04
期刊：
The Lancet. Neurology
影响因子：
--
作者：
Graus F;Titulaer MJ;Balu R;Benseler S;Bien CG;Cellucci T;Cortese I;Dale RC;Gelfand JM;Geschwind M;Glaser CA;Honnorat J;Höftberger R;Iizuka T;Irani SR;Lancaster E;Leypoldt F;Prüss H;Rae-Grant A;Reindl M;Rosenfeld MR;Rostásy K;Saiz A;Venkatesan A;Vincent A;Wandinger KP;Waters P;Dalmau J
通讯作者：
Dalmau J
影响因子：
48
作者：
Dalmau, Josep;Gleichman, Amy J.;Hughes, Ethen G.;Rossi, Jeffrey E.;Peng, Xiaoyu;Lai, Meizan;Dessain, Scott K.;Rosenfeld, Mynna R.;Balice-Gordon, Rita;Lynch, David R.
通讯作者：
Lynch, David R.
影响因子：
2.2
作者：
Adekar, Sharad P.;Jones, R. Mark;Dessain, Scott K.
通讯作者：
Dessain, Scott K.