CN114751980A - 一种用于新冠抗原检测的单抗阻断剂 - Google Patents
一种用于新冠抗原检测的单抗阻断剂 Download PDFInfo
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- CN114751980A CN114751980A CN202210429101.5A CN202210429101A CN114751980A CN 114751980 A CN114751980 A CN 114751980A CN 202210429101 A CN202210429101 A CN 202210429101A CN 114751980 A CN114751980 A CN 114751980A
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Abstract
本发明了一种针对新冠病毒S蛋白RBD蛋白的特异性单抗阻断剂。所得单抗阻断剂具有高亲和力，可以有效竞争抑制新冠病毒S蛋白RBD与ACE2的结合。假病毒中和实验显示了抗体对新冠假病毒及其突变株(Detla)假病毒均具有良好的中和活性。所述单抗阻断剂的检测下限为3.9ng/mL，线性范围为1000ng/mL～7.8ng/mL，且与新冠病毒N蛋白、流感病毒、肺炎支原体抗原及BSA、Vero细胞等均不发生交叉反应，可以极大降低试剂的假阳性，从而提高检测试剂的准确度。
Description
技术领域
本发明涉及新冠病毒检测技术领域，更具体地，涉及一种新冠病毒检测用单抗阻断剂。
背景技术
为了控制疫情的进一步发展，急需快速有效针对新冠病毒的检测手段。目前针对新冠病毒感染的诊断方法主要以对呼吸道标本进行 RT-PCR，即核酸检测为金标准，另外还有基于抗体的血清学检测、流行病学诊断和假病毒中和试验检测等。
然而，核酸检测需要相应设备，并配有专业的操作人员，且样本从采集到送检耗时长，这极大地限制了检测效率。流行病学诊断检测通量高，但需要核酸检测进行辅助检测，且检测通量低，设备要求高。假病毒中和试验检测成本高昂，且操作难度大，不适于在大流行疾病检测中推广应用。
因此，开发一种特异性强、检测灵敏的新的检测方法对该疾病预防和治疗具有重要的意义。
发明内容
针对现有技术所存在的技术问题，本发明提供了一种用于新冠抗原检测的单抗阻断剂。
具体而言，本发明首先是分离纯化了新冠病毒检测用单抗阻断剂，所述单抗阻断剂的重链可变区序列如SEQ ID NO.1所示，轻链可变区序列如SEQ ID NO.2所示；
优选地，重链可变区包括CDR1、CDR2和CDR3，其序列分别对应于 SEQ ID NO.3-5所示，轻链可变区包括CDR4、CDR5和CDR6，其序列分别对应于SEQ ID NO.6-8；
优选地，所述重链可变区的编码序列如SEQ ID NO.9所示，轻链可变区的编码序列如SEQ ID NO.10所示；
进一步地，本发明还提供了上述单抗阻断剂在制备检测新冠病毒试剂盒中的应用；优选地，所述单抗阻断剂是在间接ELISA方法中使用。
进一步地，本发明还提供了一种新冠病毒检测试剂盒，其特征在于，其所述试剂盒包括上述的单抗阻断剂；优选地，所述单抗阻断剂是在间接 ELISA方法中使用。
本发明的优点如下：本发明提供的新冠病毒单抗阻断剂是针对S蛋白 RBD蛋白的特异性抗体，所得单抗阻断剂具有高亲和力，可以有效竞争抑制新冠病毒S蛋白RBD与ACE2的结合，同时，假病毒中和实验显示了抗体对新冠假病毒及其突变株(Detla)假病毒均具有良好的中和活性。所述单抗阻断剂的检测下限为3.9ng/mL，线性范围为1000ng/mL～7.8ng/mL，且与新冠病毒N蛋白、流感病毒、肺炎支原体抗原及BSA、Vero细胞等均不发生交叉反应，可以极大降低试剂的假阳性，从而提高检测试剂的准确度。
附图说明
图1为单抗阻断剂A3-4对新冠假病毒检测灵敏度分析。
具体实施方式
下面结合具体实施例对本发明作进一步的详细说明，以使本领域的技术人员更加清楚地理解本发明。
以下各实施例，仅用于说明本发明，并不用来限制本发明的范围。基于本发明中的具体实施例，本领域普通技术人员在没有做出创造性劳动的情况下，所获得的其他所有实施例，都属于本发明的保护范围。
在本发明实施例中，若无特殊说明，所有原料组分均为本领域技术人员熟知的市售产品；在本发明实施例中，若未具体指明，所用的技术手段均为本领域技术人员所熟知的常规手段。
实施例1新冠病毒检测用单抗阻断剂的制备
采用购买自武汉华美生物工程有限公司的新冠病毒S蛋白RBD蛋白(货号CSB-DP703I)作为抗原制备单抗阻断剂。具体的实验方法：使用50μg新冠病毒S蛋白RBD蛋白对6周龄雌性BALB/c小鼠进行免疫；首次免疫后第14天采用同样方法加强免疫一次；二免后第14天进行第三次免疫注射，方法与与剂量同二免。三免后第10天对小鼠断尾采血，用间接ELISA方法监测血清抗体效价。当抗体水平达到要求后，于三免第14天后，选择效价最高的免疫小鼠，用纯的抗原液尾静脉加强免疫，3天后无菌采取小鼠脾细胞与SP2/0细胞融合制备得到杂交瘤细胞。取杂交瘤细胞株的培养上清，采用IsoStrip Mouse MonoclonalAntibody Isotyping试剂盒(Roche公司)测定抗体重链、轻链类型。
实验结果：经过三轮免疫后，最终获得了新冠病毒S蛋白RBD抗体高表达单克隆细胞株，命名为A3-4。将单克隆细胞扩增后进行腹水制备纯化抗体，用于后续亲和力测定、竞争结合实验以及假病毒(Detla)中和实验。经验证，所得单抗阻断剂具有高亲和力，可以有效竞争抑制新冠病毒S蛋白 RBD与ACE2的结合，同时，假病毒中和实验显示了抗体对新冠假病毒及其突变株(Detla)假病毒均具有良好的中和活性。且经试剂盒鉴定，所述单抗阻断剂重链可变区序列如SEQ ID NO.1所示，轻链可变区序列如SEQ ID NO.2所示。且，所述重链可变区包括CDR1、CDR2、CDR3，所述CDR1-3 的序列如SEQ ID NO.3-5所示，所述轻链可变区包括CDR4、CDR5、CDR6，所述CDR4-6的序列如SEQ ID NO.6-8所示。
实施例2单抗阻断剂A3-4与新冠病毒S蛋白RBD及其突变体(T478K,P681R和L452R)的亲和动力学分析
采用GE Biacore T200检测单抗阻断剂A3-4的亲和动力学常数，具体实验操作如仪器说明书所示。实验结果：根据检测结果，抗体亲和力数据如下表1：
表1.抗体亲和动力学分析
实验结论：本发明获得的单抗阻断剂A3-4与新冠病毒S蛋白RBD及其突变体(T478K,P681R,L452R)均具有高亲和力，尤其是其与 RBD(T478K,P681R,L452R)突变的平衡解离常数仅为4.614×10-11。
实施例3单抗阻断剂A3-4对新冠病毒S蛋白RBD及其突变体 (T478K,P681R和L452R)与ACE2结合的竞争抑制作用
利用间接ELISA方法检测单抗阻断剂A3-4对新冠病毒S蛋白RBD及其突变体(L452R，T478K)与ACE2结合的竞争抑制。检测结果如下表所示：
表2.抗体的竞争抑制作用
实验结论：本发明获得的单抗阻断剂A3-4对新冠病毒S蛋白RBD及其突变体(T478K,P681R,L452R)与ACE2结合具有明显的竞争抑制作用。
实施例4单抗阻断剂A3-4对新冠假病毒检测效果
4.1灵敏度分析
随机检测50份阴性样本，以2.1倍阴性对照的平均值作为阴性和阳性结果的判断标准，即Cut-off值。将重组RBD蛋白抗原进行2倍倍比稀释至浓度为1000ng/mL、500ng/mL～3.9ng/mL，借助间接ELISA方法，根据检测结果绘制标准曲线，确定线性范围，并根据Cut-off值确定检测下限。
4.2特异性检测
采用间接ELISE方法对SARS-CoV-2灭活病毒液、重组RBD蛋白、重组N蛋白、Vero细胞裂解液、肺炎支原体抗原、H3N2流感病毒单价原液、新生牛血清、MEM培养基、1％BSA、1％脱脂奶粉等样本进行检测。
实验结论：随机检测50份阴性样本，计算得Cut-off值为0.118；根据 Cut-off值确定检测下限为3.9ng/mL，线性范围为1000ng/mL～7.8ng/mL， R2＝0.9796(图1)。
在被检测的10份样本中，只有灭活病毒和重组RBD蛋白检测为阳性，其余样本检测结果均为阴性，均与已知背景相符。结果表明，本发明提供的单抗阻断剂，与新冠病毒N蛋白、流感病毒、肺炎支原体抗原及BSA、Vero 细胞等均不发生交叉反应。
在此有必要指出的是，以上实施例仅限于对本发明的技术方案做进一步的阐述和说明，并不是对本发明的技术方案的进一步的限制，本发明的方法仅为较佳的实施方案，并非用于限定本发明的保护范围。凡在本发明的精神和原则之内，所作的任何修改、等同替换、改进等，均应包含在本发明的保护范围之内。
SEQUENCE LISTING
<110> 厦门博昂生物技术有限公司
<120> 一种用于新冠抗原检测的单抗阻断剂
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Claims (7)
1.一种针对新冠病毒S蛋白RBD蛋白的特异性单抗阻断剂，其特征在于，所述单抗阻断剂的重链可变区包括CDR1、CDR2和CDR3，其序列分别对应于SEQ ID NO.3-5所示，轻链可变区包括CDR4、CDR5和CDR6，其序列分别对应于SEQ ID NO.6-8。
2.如权利要求1所述的单抗阻断剂，其特征在于，所述单抗阻断剂的重链可变区序列如SEQ ID NO.1所示，轻链可变区序列如SEQ ID NO.2所示。
3.如权利要求1或2所述的单抗阻断剂，其特征在于，所述单抗阻断剂的重链可变区由SEQ ID NO.9所示序列编码，轻链可变区由SEQ ID NO.10所示序列编码。
4.一种新冠病毒检测试剂盒，其特征在于，其所述试剂盒包括权利要求1-3任一项所述的单抗阻断剂。
5.如权利要求4所述的检测试剂盒，其特征在于，所述单抗阻断剂是在间接ELISA方法中使用。
6.权利要求3所述的单抗阻断剂在制备检测新冠病毒试剂盒中的应用。
7.如权利要求6所述的应用，其特征在于，所述单抗阻断剂是在间接ELISA方法中使用。
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| CN111153991A (zh) * | 2020-02-26 | 2020-05-15 | 北京博奥森生物技术有限公司 | 一种人SARS-CoV-2单克隆抗体及其制备方法和应用 |
| CN111537743A (zh) * | 2020-04-03 | 2020-08-14 | 广州医科大学附属第一医院（广州呼吸中心） | SARS-CoV-2新冠病毒抗体检测试剂盒 |
| WO2022049403A1 (en) * | 2020-09-07 | 2022-03-10 | Imperial College Innovations Limited | Sars-cov-2 antibody detection assay |
| EP3970798A1 (en) * | 2020-09-18 | 2022-03-23 | NMI Naturwissenschaftliches und Medizinisches Institut an der Universität Tübingen | Sars-cov-2-nanobodies |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111153991A (zh) * | 2020-02-26 | 2020-05-15 | 北京博奥森生物技术有限公司 | 一种人SARS-CoV-2单克隆抗体及其制备方法和应用 |
| CN111537743A (zh) * | 2020-04-03 | 2020-08-14 | 广州医科大学附属第一医院（广州呼吸中心） | SARS-CoV-2新冠病毒抗体检测试剂盒 |
| WO2022049403A1 (en) * | 2020-09-07 | 2022-03-10 | Imperial College Innovations Limited | Sars-cov-2 antibody detection assay |
| EP3970798A1 (en) * | 2020-09-18 | 2022-03-23 | NMI Naturwissenschaftliches und Medizinisches Institut an der Universität Tübingen | Sars-cov-2-nanobodies |
Non-Patent Citations (2)
| Title |
|---|
| ZEZHONG LIU,ET AL.: "RBD-Fc-based COVID-19 vaccine candidate induces highly potent SARS-CoV-2 neutralizing antibody response", 《SIGNAL TRANSDUCTION AND TARGETED THERAPY》 * |
| 夏立秋: "新型冠状病毒SARS-CoV-2研究进展", 《激光生物学报》 * |
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