CN112898416B - 新型冠状病毒np蛋白的结合蛋白及其应用 - Google Patents
新型冠状病毒np蛋白的结合蛋白及其应用 Download PDFInfo
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Abstract
本发明公开了新型冠状病毒NP蛋白的结合蛋白,其包含SEQ ID NO.1所示的CDR1、SEQ ID NO.2所示的CDR2、SEQ ID NO.3所示的CDR3以及SEQ ID NO.5所示的CDR1、SEQ ID NO.6所示的CDR2、SEQ ID NO.7所示的CDR3。本发明的结合蛋白可用于新型冠状病毒检测或新型冠状病毒感染诊断。
Description
技术领域
本发明属于细胞免疫学、分子生物学领域,涉及新型冠状病毒NP蛋白的结合蛋白及其应用。
背景技术
国际病毒分类委员会将新型冠状病毒命名为SARS-CoV-2,世界卫生组织将感染此病毒引起的肺炎称为COVID-19。此病毒传染性强,传播途径广。该病毒能迅速适应人体环境,感染后在潜伏期即具有传播能力,同时还有一些无症状感染者报道,甚至在多种动物体内也检测到病毒核酸。这些因素使得对该病毒的防控变的非常复杂,而且目前没有有效治疗药物及疫苗上市。SARS-CoV-2属于冠状病毒属,为单股正链RNA病毒,大小约30kb,与SARS-CoV相似性为79%,与蝙蝠体内分离的冠状病毒(Coronavirus,CoV)相似性最高约88%。SARS-CoV-2具有典型的冠状病毒特征,病毒包膜上有典型的棘突,形似日冕。核衣壳为螺旋对称型,主要结构蛋白是核衣壳蛋白(Nucleocapsid protein,NP),NP全长420个氨基酸。NP在病毒结构蛋白中含量最多,在宿主感染早期大量表达,且免疫原性较强,能引起宿主强烈的免疫应答。因此,NP可作为SARS-CoV-2感染血清学诊断的主要靶标抗原。
由于特异性治疗药物及有效疫苗尚未研发成功,早期诊断成为防控疫情重要的措施,早期核酸诊断及临床诊断为确诊重要依据。虽然核酸诊断速度快,受采样的质量的影响大,存在假阳性和假阴性,影响防控措施的落实。部分无症状感染者在病程晚期核酸检测也呈阴性,单凭核酸检测很容易出现漏诊。血清学诊断是检测病原感染后机体的免疫反应,持续时间长,免疫反应稳定,且免疫反应随着病程进展呈动态变化趋势。因此,血清学诊断同样也是早期诊断和感染现状评价的的重要手段。
发明内容
本发明通过提供特异性结合新型冠状病毒NP蛋白的结合蛋白(例如分离的、重组的或合成的抗体、或其片段或衍生物)来提供对于这些和其他问题的解决方案。
本发明的抗体分子可为免疫球蛋白分子的任何种类(例如IgG、IgE、IgM、IgD或IgA)或亚类。可根据所推荐的抗体分子功能选择抗体的恒定区结构域(如果存在)。例如,恒定区结构域可为人IgA、IgD、IgE、IgG或IgM结构域。尤其可使用人IgG恒定区结构域,特别是IgG1、IgG2、IgG3和IgG4。
抗体片段包括例如,例如Fab、F(ab)2、Fab’、F(ab’)2、F(ab’)3、F(v)、Fd、dAb、双抗体、小型抗体、纳米抗体片段、单个可变结构域(例如VH或VL结构域)的片段,或仅含重链或轻链结构域的片段。
本发明的抗新型冠状病毒NP蛋白的抗体,包括其片段和衍生物,可为单克隆的、多克隆的、鼠类的、嵌合的、灵长类化的、人源化的或全人源抗体。本发明的抗体可为多聚、异二聚、半二聚(hemidimeric)、单价、二价、四价、双特异性的,并且可包括单链抗体;及这些的衍生物。
在本发明的一个实施方案中,抗新型冠状病毒NP蛋白的抗体指全人源抗体。全人源抗体含具有源自人免疫球蛋白的序列(例如,得自人免疫球蛋白编码序列)的抗体多肽或免疫球蛋白可变结构域。本文中将术语“人”应用于抗体或片段(例如可变结构域)时,不含已通过将人恒定区序列移植到抗体多肽(即,用人恒定区替换非人恒定区)或通过将人V区构架序列移植到来自非人哺乳动物的免疫球蛋白可变结构域(即用人构架区替换V结构域的非人构架区)而“人源化”的来自另一个物种(例如小鼠)的抗体。
本发明的抗体含合成抗体或使用重组DNA技术产生的重组抗体,例如由噬菌体表达的抗体。还应解释为包括已通过合成编码抗体的DNA分子或确定抗体的氨基酸序列产生的抗体,所述DNA分子表达抗体蛋白、其中DNA或氨基酸序列已使用本领域可用且众所周知的合成DNA或氨基酸序列技术获得。
在某些实施方案中,本发明提供如下结合蛋白(例如抗体):其特异性结合新型冠状病毒NP蛋白,并且含一个以上的如下CDR重链(H)序列或由一个以上的如下CDR重链(H)序列构成,所述序列选自CDR-H1(SEQ ID NO.1)、CDR-H2(SEQ ID NO.2)和CDR-H3(SEQ IDNO.3)。
在进一步实施方案中,新型冠状病毒NP蛋白的结合蛋白或抗体含选自CDR-H1(SEQID NO.1)、CDR-H2(SEQ ID NO.2)和CDR-H3(SEQ ID NO.3)中的至少2个CDR,或由选自CDR-H1(SEQ ID NO.1)、CDR-H2(SEQ ID NO.2)和CDR-H3(SEQ ID NO.3)中的至少2个CDR构成。
在更进一步实施方案中,结合蛋白或抗体含所有3个如下CDR H序列或由所有3个如下CDR H序列构成,所述序列是CDR-H1(SEQ ID NO.1)、CDR-H2(SEQ ID NO.2)和CDR-H3(SEQ ID NO.3)。
在一些实施方案中,本发明提供如下结合蛋白(例如抗体):其特异性结合新型冠状病毒NP蛋白,并且含一个以上的如下CDR轻链(L)序列或由一个以上的如下CDR轻链(L)序列构成,所述序列选自CDR-L1(SEQ ID NO.5)、CDR-L2(SEQ ID NO.6)和CDR-L3(SEQ IDNO.7)。
在进一步实施方案中新型冠状病毒NP蛋白或抗体含选自CDR-L1(SEQ ID NO.5)、CDR-L2(SEQ ID NO.6)和CDR-L3(SEQ ID NO.7)中的至少2个CDR,或由选自CDR-L1(SEQ IDNO.5)、CDR-L2(SEQ ID NO.6)和CDR-L3(SEQ ID NO.7)中的至少2个CDR构成。
在更进一步实施方案中,新型冠状病毒NP蛋白的结合蛋白或抗体含所有3个如下CDR L序列或由所有3个如下CDR L序列构成,所述序列是CDR-L1(SEQ ID NO.5)、CDR-L2(SEQ ID NO.6)和CDR-L3(SEQ ID NO.7)。
在某些实施方案中,本发明提供如下结合蛋白(例如抗体):其特异性结合新型冠状病毒NP蛋白,并且含CDR-L1(SEQ ID NO.5)、CDR-L2(SEQ ID NO.6)和CDR-L3(SEQ IDNO.7),或由CDR-L1(SEQ ID NO.5)、CDR-L2(SEQ ID NO.6)和CDR-L3(SEQ ID NO.7)构成,并且其中所述结合蛋白进一步含CDR-H1(SEQ ID NO.1)、CDR-H2(SEQ ID NO.2)和CDR-H3(SEQID NO.3),或由CDR-H1(SEQ ID NO.1)、CDR-H2(SEQ ID NO.2)和CDR-H3(SEQ ID NO.3)构成。
在某些实施方案中,本发明提供特异性结合新型冠状病毒NP蛋白的结合蛋白(例如抗体),其中所述新型冠状病毒NP蛋白的结合蛋白或抗体含SEQ ID NO.4的VH序列,或由SEQ ID NO.8的VH序列构成。
在某些实施方案中,本发明提供特异性结合新型冠状病毒NP蛋白的结合蛋白(例如抗体),其中所述新型冠状病毒NP蛋白的结合蛋白或抗体含SEQ ID NO.8的VL序列,或由SEQ ID NO.8的VL序列构成。
在某些实施方案中,本发明提供特异性结合新型冠状病毒NP蛋白的结合蛋白(例如抗体),其中所述新型冠状病毒NP蛋白的结合蛋白或抗体含SEQ ID NO.8的VL序列和SEQID NO.4的VH序列,或由SEQ ID NO.8的VL序列和SEQ ID NO.4的VH序列构成。
本发明的新型冠状病毒NP蛋白的结合蛋白(例如抗体)可经PEG化。其中抗体可在重链、轻链或两条链上经PEG化。
本发明还提供了编码前面所述的结合蛋白的分离的、重组的和/或合成的DNA分子。
本发明的DNA序列可含例如通过化学处理产生的合成DNA、cDNA、基因组DNA或其任何组合。
在本发明的具体实施方案中,所述DNA分子包含SEQ ID NO.17和SEQ ID NO.18所示的氨基酸序列。
本发明还提供了含有前面所述的DNA分子的载体。
本发明还提供含本发明的DNA序列或载体的宿主细胞。在某些方面,本发明涉及用于产生前面所述的结合蛋白(例如抗体)的方法,其包括在适合于由宿主细胞产生结合蛋白的条件下,培养含上述任何载体的宿主细胞。在一些实施方案中,该方法包括从宿主细胞培养物中回收结合蛋白。
宿主细胞可为例如原核细胞例如大肠杆菌,或其他微生物细胞,或真核细胞包括但不限于哺乳动物细胞,例如人、小鼠、猴、兔、山羊、仓鼠或大鼠细胞,昆虫细胞,禽类细胞,植物细胞和低等真核细胞例如真菌细胞。
在本发明的一些实施方案中,用于实践本发明的宿主细胞可为例如(1)细菌细胞例如大肠杆菌、新月柄杆菌(Caulobacter crescentus)、链球菌属(Streptococci)、葡萄球菌属(Staphylococci)、链霉菌属(Streptomyces)物种和枯草芽孢杆菌(Bacillussubtilis)细胞、和鼠伤寒沙门氏菌(Salmonella typhimurium);(2)真菌细胞和曲霉菌属(Aspergillus)细胞、酵母细胞,例如酿酒酵母(Saccharomyces cerevisiae)、栗酒裂殖酵母(Schizosaccharomyces pombe)、巴斯德毕赤酵母(Pichia pastoris)、甲醇毕赤酵母(Pichia methanolica)、其他毕赤酵母属物种、乳酸克鲁维氏酵母(K.lactis),(3)昆虫细胞系,例如来自草地贪夜蛾(Spodoptera frugiperda)的细胞(如Sf9和Sf21细胞系,以及expresSFTM细胞(Protein Sciences Corp.,Meriden,CT,USA))、果蝇属S2细胞,和粉纹叶蛾(Trichoplusia ni)High Cells(Invitrogen,Carlsbad,CA,USA);(4)哺乳动物细胞,或(5)植物细胞。
一般的哺乳动物细胞包括COS1和COS7细胞、中国仓鼠卵巢(CHO)细胞、NSO骨髓瘤细胞、NIH3T3细胞、293细胞、HΕPG2细胞、HeLa细胞、C127、3T3、BHK、Bowes黑素瘤细胞、L细胞、MDCK、HEK293、WI38、鼠类ES细胞系(例如,来自品系129/SV、C57/BL6、DBA-1、129/SVJ)、K562、Jurkat细胞和BW5147。
在某些实施方案中,本发明的结合蛋白、或核酸经可检测标记进行标记,所述可检测标记可为放射性同位素、酶、染料或生物素。
在另外其他的实施方案中,本发明的抗体与可为标记部分的显像剂缀合。标记剂可为生物素、荧光或发光部分、放射性部分、组氨酸标签或肽标签。
本发明还涉及本文所述结合蛋白(例如抗体)、以及编码其的核酸序列的序列变体。本发明的序列变体优选与本发明的多肽或编码其的核酸序列享有至少90%、91%、92%、93%或94%同一性。序列变体在氨基酸或核酸水平上更优选享有至少95%、96%、97%或98%同一性。序列变体与本发明的多肽或编码其的核酸序列最优选享有至少99%、99.5%、99.9%或更多同一性。
在某些实施方案中,本发明的抗新冠状病毒NP蛋白的抗体多肽是例如dAb、Fab、Fab’、scFv、Fv、二硫键合的Fv,或含单免疫球蛋白可变结构域,例如VH或VL结构域,其对于新冠状病毒NP蛋白的结合特异且是单价的。本发明的某些结合蛋白多肽含本发明的1个、2个或更多本发明的CDR和备选的支架或通用构架序列。
在某些实施方案中,本发明的结合蛋白或抗体含选自可变重链(VH)和可变轻链(VL)的单可变结构域。
本发明还提供了含本发明前面所述的结合蛋白的组合物。
所述组合物还可进一步含第二种结合蛋白,所述第二种结合蛋白包含重链可变区CDR1、重链可变区CDR2、重链可变区CDR3、轻链可变区CDR1、轻链可变区CDR2、轻链可变区CDR3;其中,
重链可变区CDR1含有SEQ ID NO.9所示的氨基酸序列;
重链可变区CDR2含有SEQ ID NO.10所示的氨基酸序列;
重链可变区CDR3含有SEQ ID NO.11所示的氨基酸序列;
轻链可变区CDR1含有SEQ ID NO.13所示的氨基酸序列;
轻链可变区CDR2含有SEQ ID NO.14所示的氨基酸序列;
轻链可变区CDR3含有SEQ ID NO.15所示的氨基酸序列;
更优选地,所述第二种结合蛋白包含重链可变区、轻链可变区;其中,重链可变区含有SEQ ID NO.12所示的氨基酸序列,轻链可变区含有SEQ ID NO.16所示的氨基酸序列。
所述第二种结合蛋白的实例是单克隆抗体。
所述组合物还可进一步含有另外的生物活性剂或诊断剂。
本发明的抗体或含其的组合物可单独或作为试剂盒的部分与标签和施用说明书一起包括在容器、包装或分配器中。
所述诊断剂包括可检测物质,可检测物质的例子包括各种酶、辅基、荧光材料、发光材料、生物发光材料、放射性核素、正电子发射金属(用于在正电子发射断层摄影术中使用)、和非放射性顺磁金属离子。关于可与抗体缀合用于用作诊断剂的金属离子,一般参见US4,741,900。合适的酶包括辣根过氧化物酶、碱性磷酸酶、β-半乳糖苷酶、或乙酰胆碱酯酶;合适的辅基包括链霉亲和素、抗生物素蛋白和生物素;合适的荧光材料包括伞形酮、荧光素、异硫氰酸荧光素、罗丹明、二氯三嗪基胺荧光素、丹酰氯和藻红蛋白;合适的发光材料包括鲁米诺;合适的生物发光材料包括萤光素酶、萤光素和水母素;放射性同位素,例如125I、131I、111In和90Y、Lu177、铋213、锎252、铱192和钨188/铼188、211砹、99Tc。
术语“聚乙二醇化”、“聚乙二醇”或“PEG”包括聚亚烷基二醇化合物或其衍生物,连同或不连同偶联剂或者用偶联或活化部分(例如用硫醇、三氟甲磺酸盐(triflate)、三氟乙磺酸盐(tresylate)、氮丙啶、环氧乙烷,或优选用马来酰亚胺部分(例如PEG-马来酰亚胺))的衍生化。其他合适的聚亚烷基二醇化合物包括但不限于,马来酰亚胺单甲氧基PEG、活化PEG聚丙二醇、以及下述类型的带电或中性聚合物:右旋糖、多聚乙酰神经氨酸、或其他基于碳水化合物的聚合物、氨基酸聚合物、以及生物素和其他亲和剂衍生物。
本发明还提供了制备本发明的结合蛋白(例如抗体)的方法,包括用永生性细胞株、人工合成、重组表达或噬菌体展示技术等方法来制备。在具体实施方案中,本发明的方法包括培养前面所述的宿主细胞的步骤。
本发明提供了一种应用,其包含以下任一项所述的应用:
(1)前面所述的结合蛋白在制备新型冠状病毒检测产品中的应用;
(2)前面所述的结合蛋白在制备新型冠状病毒感染诊断产品中的应用;
(3)前面所述的组合物在制备新型冠状病毒的检测产品中的应用。
所述检测产品或诊断产品包括前面所述的试剂盒。
附图说明
图1显示本发明的重组SARS-CoV2 NP蛋白的SDS-PAGE图;
图2显示利用间接ELISA检测抗体滴度的结果图;
图3显示利用WB检测抗体与抗原结合的结果图;
图4显示利用SPR检测JS01的亲和活性结果图;
图5显示利用SPR检测JS02的亲和活性结果图;
图6显示利用SPR检测JS03的亲和活性结果图;
图7显示利用SPR检测JS04的亲和活性结果图;
图8显示利用SPR检测JS05的亲和活性结果图;
图9显示利用SPR检测JS06的亲和活性结果图;
图10显示利用SPR检测JS07的亲和活性结果图;
图11显示利用SPR检测JS08的亲和活性结果图;
图12显示利用SPR检测JS09的亲和活性结果图;
图13显示利用SPR检测JS10的亲和活性结果图;
图14显示利用SPR检测JS11的亲和活性结果图;
图15显示利用SPR检测JS12的亲和活性结果图;
图16显示利用SPR检测JS13的亲和活性结果图;
图17显示利用SPR检测JS14的亲和活性结果图;
图18显示利用SPR检测JS15的亲和活性结果图;
图19显示利用SPR检测JS16的亲和活性结果图;
图20显示利用双抗体夹心法检测抗体包被浓度的结果图;
图21显示利用双抗体夹心法检测抗体检测灵敏度的结果图;
图22显示本发明的抗原检测层析条的检测效果图。
具体实施方式
下面通过附图和实施例进一步说明本发明。应该理解的是,本发明的实施例是用于说明本发明而不是对本发明的限制。根据本发明的实质对本发明进行的简单改进都属于本发明要求保护的范围。
实施例1抗体筛选
一、重组SARS-CoV2核蛋白(NP)表达
1.1主要试剂
SARS-CoV2 NP基因序列(GenBank序列号:MT066176.1)以及相关引物合成和测序均由通用生物系统(安徽)有限公司完成;E.coli DH5α,BL21(DE3)感受态细胞购自通用生物系统(安徽)有限公司;BamHⅠ和NotⅠ核酸内切酶购自New England Biolabs(NEB)公司;EXTaq酶购自TaKaRa公司;HRP标记抗人Fc抗体购自Sigma公司;其他化学试剂均为国产分析纯试剂;感染2019-nCoV患者血清由本中心收集及保存。
1.2原核表达质粒构建
设计NP基因原核表达引物,上游引物带BamHⅠ酶切位点,下游引物带NotⅠ酶切位点。引物序列为:Cov2-NP-F:CGGGATCCTCTGATAATGGACCCCAAAATC;Cov2-NP-R:ATAAGAATGCGGCCGCAGGCCTGAGTTGAGTCAGCAC。使用EX Taq酶扩增NP基因,PCR反应程序为:94℃3min;94℃30s,58℃30s,72℃80s,共30个循环;72℃10min。PCR产物经胶回收约1 300bp目的片段,然后经BamHⅠ和NotⅠ双酶切后连接pET28a载体,转化E.coli DH5α感受态细胞。次日挑选单菌落测序正确后,提质粒转化原核表达菌E.coli BL21(DE3)感受态细胞。
1.3 NP表达及纯化
将NP表达菌培养至OD600=0.6后,加入终浓度0.5mmol/L的IPTG,16℃诱导6h,收集菌体,超声破碎后离心收集包涵体。将包涵体溶于8mol/L尿素中,然后用镍柱亲和层析纯化包涵体蛋白。纯化后梯度减少尿素含量,使蛋白透析复性至PBS中,最后经SDS-PAGE检测蛋白表达及纯化效果。小量表达成功以后即上100L发酵罐进行大量发酵,发酵培养基为TB培养基(1%甘油),发酵参数为280rpm,通气比为0.5vvm(15L/min),pH控制在6.8~7.2,罐压0.06MPa~0.1MPa,发酵温度为16℃,培养24h。
SDS-PAGE结果显示:NP全长加上载体中的His tag及其他附带氨基酸,预计蛋白相对分子质量约50×103。表达菌经IPTG诱导后,发现在50×103左右出现明显的条带,与预计分子量大小一致(图1A)。包涵体溶解后,过镍柱纯化,在150mmol/L咪唑时有明显的洗脱峰。蛋白经透析复性后,经SDS-PAGE发现在同样的位置出现单一蛋白条带(图1B)。此说明NP被成功诱导表达并且纯化后纯度较高。注:图中M:蛋白Makers;1:未诱导的pET28a-NP表达菌;2:IPTG诱导后pET28a-NP重组表达菌;3:纯化后重组核衣壳蛋白。
二、噬菌体文库构建
1、采集COVID-19患者恢复期外周血,从外周血中分离单个核细胞(PBMC)
经过知情同意,采集5位COVID-19确诊患者出院前外周血各20ml。使用GE的Ficoll-Paque PLUS,经密度梯度离心法,分离20ml肝素抗凝血中的单个核细胞(PBMC)。
2、PBMC中RNA的提取和cDNA的合成
使用QIAGEN的RNeasy Mini Kit提取PBMC细胞RNA,然后使用罗氏公司的第一链合成试剂盒(Transcriptor First Strand cDNA Synthesis Kit,Roche,Cat No.:04896866001)将RNA反转录成cDNA。
3、PCR扩增VK,VL和VH(EX Taq,Takara,Cat No.:DRR001A)
(1)扩增VK&VL体系如表1所示。
表1扩增VK&VL体系
溶液或组分 | 体积(μL) |
cDNA | 1 |
EX Buffer(10x) | 5 |
dNTPs(10mM each) | 4 |
P1(10μM) | 2 |
P2(10μM) | 2 |
EX Taq 1U/μl | 0.3 |
dH<sub>2</sub>O | 35.7 |
(2)扩增重链Fd段体系如表2所示。
表2扩增重链Fd段体系
溶液或组分 | 体积(μL) |
cDNA | 2 |
EX Buffer(10x) | 10 |
dNTPs(10mM each) | 8 |
P1(10μM) | 2 |
P2(10μM) | 2 |
EX Taq 1U/μl | 0.6 |
dH<sub>2</sub>O | 75.4 |
(3)反应程序如表3所示。
表3反应程序
PCR产物过2%琼脂糖凝胶电泳,回收750bp左右的片段。
4、轻链克隆(将VK和VL克隆入pComb3H载体)
VK和VL经XbaⅠ和SacⅠ酶切后与同样经XbaⅠ和SacⅠ酶切的pComb3H载体连接后,回收连接产物,然后电转XL1-Blue感受态细胞。
电击菌液涂15cm大平皿,次日刮菌,提质粒即为轻链库。此时重组质粒为pComb3H-VK和pComb3H-VL。
5、重链克隆(将VH基因克隆入pComb3H-VK和pComb3H-VL轻链库中)
将轻链库pComb3-L和Fd片段分别经XhoI和SpeI双酶切,与同样经XhoI和SpeI双酶切的pComb3H-VK和pComb3H-VL连接,然后电转即得到抗体文库。
6、抗体库的包装
(1)从-80℃冰箱取出抗体库,冰上融化后取1ml加入10ml A+(20μg/ml)2YT培养基中,37℃200rpm摇1小时;
(2)加100ml A+(100μg/ml),T+(20μg/ml)2YT培养基,200rpm摇1小时;
(3)加1012pfu的VCSM13辅助噬菌体,37℃静置20min,200rpm摇2小时;
(4)加终浓度70μg/ml卡那30℃200rpm摇过夜;
(5)次日6000rpm离心20min,倒出上清,加入4%PEG8000(4g)和3%NaCl(3g),混匀,置于冰上30min以上;
(6)分装于50ml离心管中9000rpm离心25min,弃去上清,控干,沉淀用1ml PBS重悬即为包装文库。
三、噬菌体文库筛选
1、将重组SARS-CoV2核蛋白(NP)包被于免疫管中,按50μg/管包被3管,于4℃放置过夜,次日用2%脱脂牛奶封闭免疫管1h。
2、向免疫管中加1.75ml含2%脱脂牛奶的PBS和250μl噬菌体文库,37℃摇1h,再37℃静置1h。
3、倒去噬菌体文库,用PBST洗20次,每次摇5min。
4、用1ml pH=2.2的Gly-HCl洗脱免疫管,室温静置5min,再37℃摇5min,然后吸至1.5ml EP管中,加入57μl 2M Tris中和至pH=7。
5、将洗脱液转移至一个新的50ml离心管中,立即加入10ml OD=1的新鲜XL1-Blue,混匀后37℃孵育30min,加入10ml 2YT(Amp 100μg/ml,Tet 20μg/ml)。
6、取10μl菌液用来测洗脱库容量,剩下20ml培养基倒入500ml三角瓶,230rpm摇1h。
7、加入130ml 2YT(Amp 100ug/ml,Tet 20μg/ml),230rpm摇1h。
8、加入MOI=20的辅助噬菌体,37℃静置孵育30min。
9、3000g 10min离心,重悬沉淀至150ml 2YT(Amp 100μg/ml,Tet 20μg/ml)中,37℃,230rpm摇2h。
10、加入110μl 70mg/ml卡那霉素,30℃230rpm过夜。次日再加1/5体积的PEG-NaCl(40ml),混匀后冰浴至少1h,然后10000g 4℃离心20min,沉淀重悬于2-3ml PBS中,瞬时离心去除杂菌,过0.45μm滤器后用于下一轮筛选。
11、重复上述筛选步骤3次,以达到对噬菌体文库富集筛选的目的。
12、第三轮富集完以后,挑选2*96个克隆。经IPTG诱导后,次日进行ELSA检测。
四、ELISA检测2*96个克隆的结合特异性
1、分别包被2块抗人Fab抗体(1:3000)和2块NP蛋白(2μg/ml),于4℃包被过夜。
2、次日用3%脱脂牛奶封闭1h,然后加入50μl诱导上清和50μl脱脂牛奶,37℃孵育1h,PBST洗涤。
3、4块板均加入HRP标记的抗人Fab抗体(1:3000),37℃孵育1h,PBST洗涤后,TMB显色。
经筛选共获得178株能与NP特异性结合的噬菌体抗体片段,该片段为人源的Fab段,包括轻链全长及重链的Fd段。将178个单菌落扩增后送测序,共获得159株轻重链齐全的合格序列。
实施例2全抗体的表达及相关功能验证
从获得的159株抗体中最终选定16株抗体用于全抗体的表达及相关功能验证,将此16株抗体命名为JS01~JS16。
其中,JS03抗体序列如下所示:
重链可变区CDR1的氨基酸序列如SEQ ID NO.1所示;
重链可变区CDR2的氨基酸序列如SEQ ID NO.2所示;
重链可变区CDR3的氨基酸序列如SEQ ID NO.3所示;
轻链可变区CDR1的氨基酸序列如SEQ ID NO.5所示;
轻链可变区CDR2的氨基酸序列如SEQ ID NO.6所示;
轻链可变区CDR3的氨基酸序列如SEQ ID NO.7所示;
重链可变区的氨基酸序列如SEQ ID NO.4所示,核酸序列如SEQ ID NO.17所示;轻链可变区的氨基酸序列如SEQ ID NO.8所示,核酸序列如SEQ ID NO.18所示。
JS08抗体序列如下所示:
重链可变区CDR1的氨基酸序列如SEQ ID NO.9所示;
重链可变区CDR2的氨基酸序列如SEQ ID NO.10所示;
重链可变区CDR3的氨基酸序列如SEQ ID NO.11所示;
轻链可变区CDR1的氨基酸序列如SEQ ID NO.13所示;
轻链可变区CDR2的氨基酸序列如SEQ ID NO.14所示;
轻链可变区CDR3的氨基酸序列如SEQ ID NO.15所示。
重链可变区的氨基酸序列如SEQ ID NO.12所示;轻链可变区的氨基酸序列如SEQID NO.16所示。
1、全抗体表达
将上述16株人源抗体,构建成IgG形式人源全分子抗体,并在293F细胞中表达,用Protein A纯化后备用。
2、ELISA检测16株抗体与重组NP结合特异性
将重组NP用PBS按1μg/ml浓度包被ELISA板,将所有抗体浓度稀释到1mg/ml,然后从1:10000开始倍比稀释,37℃孵育30min。然后PBST洗3次,加入HRP标记的抗人IgG(1:5000),37℃孵育30min后PBST洗3次,然后TMB显色,终止后读取OD450吸光度值。
采用间接ELISA检测16株NP抗体的稀释滴度,阴性对照的平均OD值为0.119,标准差为0.132,因此将cutoff值定义为由此判断16株抗体的检测滴度为1:80000~1:1280000之间(图2)。
3、16株抗体与纯化NP的Western Blot结果
将1μg重组NP经SDS-PAGE电泳后,转印至PVDF膜中,用上述16种抗体(0.5μg/ml)37℃孵育1h,用PBST洗涤3次,然后用HRP标记的抗人IgG(1:5000)孵育30min,PBST洗涤3次,然后用DAB之间在膜上显色。
WB实验结果显示16株抗体均能特异性结合重组表达的核蛋白(NP),并在50kDa处出现明显的显色带,此提示该组抗体全部为线性表位的抗体(图3)。
4、抗体亲和活性检测
抗体亲和力测定由Biacore T200工作站完成,具体按以下步骤进行:首先使用氨基偶联活化剂NHS和EDC以10μl/min 300s活化CM5芯片,然后用10mM醋酸钠缓冲液(pH5.5)将重组表达的SARS-CoV-2NP稀释到1ug/mL,以10μl/min流经芯片30s使响应值(Responseunits,RUs)达到600左右,最后设置10μl/min、420s,进样乙醇胺对芯片表面剩余活化位点进行封闭。系列稀释的抗体,在25℃时以30μl/min流速依次进样,每测定一次浓度以后用pH2.0的甘氨酸-盐酸再生CM5芯片,然后进行下个浓度检测。实验结束后,利用Biacore T200Evaluation Software软件全局拟合曲线来获得结合亲和力。
实验结果如图4-19所示,JS01-JS16可高效结合SARS-CoV-2NP蛋白,亲和活性相关参数见表4。
表4抗体亲和力参数
抗体名称 | 配体偶联量 | ka(1/Ms) | Kd(1/s) | KD(M) |
JS01 | 102RU | 2.18E+06 | 4.79E-04 | 2.20E-10 |
JS02 | 162RU | 9.12E+05 | 5.83E-05 | 6.39E-11 |
JS03 | 102RU | 8.97E+05 | 2.12E-04 | 2.36E-10 |
JS04 | 162RU | 7.15E+04 | 1.91E-04 | 2.67E-09 |
JS05 | 162RU | 5.94E+05 | 2.43E-04 | 4.09E-10 |
JS06 | 136RU | 1.61E+05 | 0.002576 | 1.61E-08 |
JS07 | 110RU | 1.44E+06 | 1.78E-04 | 1.24E-10 |
JS08 | 162RU | 3.03E+04 | 5.77E-06 | 1.90E-10 |
JS09 | 129RU | 6.89E+05 | 4.79E-05 | 6.94E-11 |
JS10 | 136RU | 1.47E+06 | 2.99E-04 | 2.04E-10 |
JS11 | 110RU | 1.93E+05 | 5.62E-05 | 2.92E-10 |
JS12 | 110RU | 4.88E+05 | 7.33E-05 | 1.50E-10 |
JS13 | 110RU | 8.05E+05 | 9.66E-05 | 1.20E-10 |
JS14 | 136RU | 1.24E+06 | 2.21E-04 | 1.78E-10 |
JS15 | 110RU | 7.72E+04 | 1.22E-04 | 1.58E-09 |
JS16 | 136RU | 2.68E+05 | 4.01E-05 | 1.50E-10 |
5、抗体配对实验
5.1抗体包被浓度的确定
(1)将100μl JS12抗体从5μg/ml开始倍比稀释到0.0024μg/ml共稀释12个稀释度,然后包被于ELISA板中。4℃包被过夜,次日用1%BSA封闭2h,PBST洗3次。
(2)向每个包被浓度的第一孔加入50ng的重组NP,然后倍比稀释到0.39ng/孔,共8个稀释度,37℃孵育1h,PBST洗3次。
(3)加入1:1000稀释HRP标记的JS08,37℃孵育1h,PBST洗3次,TMB显色后读取OD450nm吸光度值。
由图20的曲线可以看出,包被抗体的量对检测灵敏性有影响,从5μg/ml到0.00245μg/ml的包被量看来,影响不大,检测NP抗原的灵敏性看小于3.9ng/ml。因此,后续配对实验,我们均选择2μg/ml浓度作为抗体包被量,用1:4000作为酶标抗体的稀释度。
5.2双抗夹心法检测NP
(1)将16株NP抗体JS01~JS16按2μg/ml浓度包被ELISA板,于4℃包被过夜,次日用1%BSA封闭2h,PBST洗3次。
(2)加入0.1μg/ml重组NP蛋白,然后倍比稀释至0.78ng/ml,共8个稀释度,37℃孵育1h,PBST洗3次。
(3)加入HRP标记的JS08(1:1000),37℃孵育1h,PBST洗3次,TMB显色,读取OD450nm吸光度值。
由图21可见,酶标的JS08不能与JS06,JS11和JS08自身配对,而与其他13株NP抗体均可配对用于双抗夹心法检测NP。其中JS08与JS16配对效果最好,其检测限可达0.78ng/ml以下,其他配对抗体检测限在12.5-1.56ng/ml之间。
6、双抗体夹心免疫层析法检测重组NP的灵敏度
将抗JS08单抗包被在硝酸纤维素膜上形成T线,抗人IgG抗体标记至C线处。NP蛋白系列稀释后加50μL于样本孔中,样本孔下的结合垫上的标有色微球的JS01~JS16抗体与NP形成免疫复合物,然后通过层析作用迁移至T线处,与此处标记抗体结合固定,形成有色T线。多余的人源单抗再继续迁移至C线处,与抗人抗体结合,形成C线。以此来测定对NP的结合灵敏度。
采用有色微球标记JS08抗体与13株抗体分别配对,制成抗原检测层析条。用2ng/ml的重组NP验证试纸条,从结果可见全部层析条均可见明显的检测T线,而质控C线也非常明显(图22)。这说明13对抗体组合均可用于检测新冠状病毒的核蛋白,且检测限小于2ng/ml。
虽然以上仅描述了本发明的具体实施方式范例,但是本领域的技术人员应当理解,这些仅是举例说明,本发明的保护范围是由所附权利要求书限定的。本领域的技术人员在不背离本发明的原理和实质的前提下,可以对这些实施方式做出多种变更或修改,但这些变更或修改均落入本发明的保护范围。
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<110> 江苏省疾病预防控制中心(江苏省公共卫生研究院)
<120> 新型冠状病毒NP蛋白的结合蛋白及其应用
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Claims (11)
1.新型冠状病毒NP蛋白的结合蛋白,其包含SEQ ID NO.1所示的重链可变区CDR1、SEQID NO.2所示的重链可变区CDR2、SEQ ID NO.3所示的重链可变区CDR3、SEQ ID NO.5所示的轻链可变区CDR1、SEQ ID NO.6所示的轻链可变区CDR2和SEQ ID NO.7所示的轻链可变区CDR3。
2.如权利要求1所述的结合蛋白,其包含SEQ ID NO.4所示的重链可变区以及SEQ IDNO.8所示的轻链可变区。
3.分离的、重组的或合成的DNA分子,其编码权利要求1或2所述的结合蛋白。
4.如权利要求3所述的DNA分子,其包含SEQ ID NO.17和18所示的序列。
5.载体,其包含权利要求3或4所述的DNA分子。
6.宿主细胞,其含权利要求5所述的载体。
7.用于产生新型冠状病毒NP蛋白的结合蛋白的方法,其包括在适合于宿主细胞产生新型冠状病毒NP蛋白的结合蛋白的条件下,培养权利要求6所述的宿主细胞。
8.一种组合物或试剂盒,其含有权利要求1或2所述的结合蛋白。
9.如权利要求8所述的组合物或试剂盒,所述组合物或试剂盒还包含第二种结合蛋白,所述第二种结合蛋白包含重链可变区CDR1、重链可变区CDR2、重链可变区CDR3、轻链可变区CDR1、轻链可变区CDR2和轻链可变区CDR3;其中,
重链可变区CDR1含有SEQ ID NO.9所示的氨基酸序列;
重链可变区CDR2含有SEQ ID NO.10所示的氨基酸序列;
重链可变区CDR3含有SEQ ID NO.11所示的氨基酸序列;
轻链可变区CDR1含有SEQ ID NO.13所示的氨基酸序列;
轻链可变区CDR2含有SEQ ID NO.14所示的氨基酸序列;
轻链可变区CDR3含有SEQ ID NO.15所示的氨基酸序列。
10.如权利要求9所述的组合物或试剂盒,所述第二种结合蛋白包含重链可变区和轻链可变区;其中,重链可变区含有SEQ ID NO.12所示的氨基酸序列,轻链可变区含有SEQ IDNO.16所示的氨基酸序列。
11.一种应用,其包含以下任一项所述的应用:
(1)权利要求1或2所述的结合蛋白在制备新型冠状病毒检测产品中的应用;
(2)权利要求1或2所述的结合蛋白在制备新型冠状病毒感染诊断产品中的应用;
(3)权利要求8-10中任一项所述的组合物在制备检测新型冠状病毒的产品中的应用。
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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 |
CN111074008A (zh) * | 2020-02-28 | 2020-04-28 | 南京申基医药科技有限公司 | 一种可提高准确率的covid-19新型冠状病毒核酸检测方法 |
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Non-Patent Citations (1)
Title |
---|
Current practice and potential strategy in diagnosing COVID-19;D-Y Wan,等;《Eur Rev Med Pharmacol Sci》;20200430;第24卷(第8期);第4548-4553页 * |
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