Single-Cell Sorting of HBsAg-Binding Memory B Cells from Human Peripheral Blood Mononuclear Cells and Antibody Cloning.
Single-Cell Sorting of HBsAg-Binding Memory B Cells from Human Peripheral Blood Mononuclear Cells and Antibody Cloning.
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人外周血单核细胞中 HBsAg 结合记忆 B 细胞的单细胞分选和抗体克隆
DOI：
10.1016/j.xpro.2020.100129
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发表时间：
2020-12-18
期刊：
影响因子：
--
通讯作者：
Wang Q
中科院分区：
文献类型：
--
作者：
Zhou Y;Liu Z;Wang Z;Zhang Q;Mayer CT;Schoofs T;Nussenzweig MC;de Jong YP;Wang Q
具有天然配对的重链和轻链的人抗体的分离对于理解人抗体免疫应答至关重要。在这里，我们提出了一个方案，从分选的单个人记忆B细胞识别乙型肝炎病毒（HBV）S抗原（HBsAg）的抗体克隆。针对HBsAg的双荧光染料标记策略允许提高分选特异性，而非相关蛋白质染色允许排除非特异性B细胞。该方案也可广泛适用于其他抗原。有关本方案使用和执行的完整详细信息，请参阅。将识别HBsAg的人记忆B细胞单细胞分选到96孔板中，使用双荧光染料标记策略来提高分选特异性。天然配对抗体的克隆对于研究人抗体应答是至关重要的具有天然配对重链和轻链的人抗体的分离对于理解人抗体应答是至关重要的。抗体免疫反应在这里，我们提出了一个方案，从分选的单个人记忆B细胞识别乙型肝炎病毒（HBV）S抗原（HBsAg）的抗体克隆。针对HBsAg的双荧光染料标记策略允许提高分选特异性，而非相关蛋白质染色允许排除非特异性B细胞。该方案也可广泛适用于其他抗原。
The isolation of human antibodies with naturally paired heavy and light chains is crucial for understanding the human antibody immune response. Here, we present a protocol for antibody cloning from the sorted single human memory B cells recognizing hepatitis B virus (HBV) S antigen (HBsAg). A two-fluorescent-dye labeling strategy against HBsAg allows for an improved sorting specificity, while non-relevant protein staining allows for the exclusion of non-specific B cells. This protocol could also be widely adapted for other antigens. For complete details on the use and execution of this protocol, please refer to. Human memory B cells recognizing HBsAg were single-cell sorted into 96-well plates A two-fluorescent-dye labeling strategy was used to improve the sorting specificity Three nested PCRs were performed using reverse-transcribed cDNA of sorted single cells Cloning of naturally paired antibody is critical for studying the human antibody response The isolation of human antibodies with naturally paired heavy and light chains is crucial for understanding the human antibody immune response. Here, we present a protocol for antibody cloning from the sorted single human memory B cells recognizing hepatitis B virus (HBV) S antigen (HBsAg). A two-fluorescent-dye labeling strategy against HBsAg allows for an improved sorting specificity, while non-relevant protein staining allows for the exclusion of non-specific B cells. This protocol could also be widely adapted for other antigens.
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影响因子：
14.9
作者：
Ye J;Ma N;Madden TL;Ostell JM
通讯作者：
Ostell JM
影响因子：
14.8
作者：
von Boehmer, Lotta;Liu, Cassie;Nussenzweig, Michel C.
通讯作者：
Nussenzweig, Michel C.
影响因子：
14.9
作者：
Brochet X;Lefranc MP;Giudicelli V
通讯作者：
Giudicelli V
人类广泛中和乙型肝炎病毒 HBsAg 的抗体与不同表位的组合可抑制逃逸突变
影响因子：
30.3
作者：
Wang Q;Michailidis E;Yu Y;Wang Z;Hurley AM;Oren DA;Mayer CT;Gazumyan A;Liu Z;Zhou Y;Schoofs T;Yao KH;Nieke JP;Wu J;Jiang Q;Zou C;Kabbani M;Quirk C;Oliveira T;Chhosphel K;Zhang Q;Schneider WM;Jahan C;Ying T;Horowitz J;Caskey M;Jankovic M;Robbiani DF;Wen Y;de Jong YP;Rice CM;Nussenzweig MC
通讯作者：
Nussenzweig MC