CN117264061B - 一种识别uPAR的纳米抗体及其应用 - Google Patents
一种识别uPAR的纳米抗体及其应用 Download PDFInfo
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Abstract
本发明提供了一种识别uPAR的纳米抗体及其应用,该纳米抗体或其抗原结合片段对uPAR蛋白具有较高的亲和力,相比于传统抗体技术具有特异性强、亲和力高、分子量小、可溶性高、稳定性强、结构简单、基因工程易改造等优势。uPAR在衰老、部分肿瘤及炎性疾病病理过程中被广泛诱导,其可作为衰老细胞、肿瘤细胞和炎性组织表面标志物,因此uPAR纳米抗体还可作为检测体内衰老细胞、肿瘤细胞及炎性组织的探针,当其偶联成像剂时还可进行定位成像。同理在衰老、肿瘤等靶向药物的设计中,uPAR纳米抗体亦可作为药物靶标,可准确识别并结合uPAR从而定位衰老细胞和肿瘤细胞进行靶向作用或给药。
Description
技术领域
本发明涉及抗体技术领域,更具体地,涉及一种识别uPAR的纳米抗体及其应用。
背景技术
尿激酶型纤溶酶原激活物(uPA)受体(uPAR)是一种GPI锚定的细胞膜受体,通过共价键与细胞膜外层相连,由三个同源结构域(D1,D2,D3)组成。D1区结合uPA,D3区通过糖基磷脂酰肌醇尾部将uPAR锚定在细胞膜表面。uPAR主要功能是激活细胞表面的尿激酶(uPA)蛋白水解活性。完整的uPAR以高亲和力(1nM)结合uPA和uPA酶原,当uPA酶原结合在uPAR上时可以被纤溶酶激活成为具有蛋白水解活性的uPA,从而在纤维蛋白溶解、伤口愈合或肿瘤发生过程中促进细胞外基质的降解。一部分uPAR在配体结合后被蛋白水解切割,生成血清可溶性uPAR(suPAR)。
研究表明uPAR与人类炎症性及感染性疾病有关,感染细菌或人类免疫缺陷病毒-1(HIV-1)时,白细胞表面表达的uPAR及血清中的suPAR数量增加,suPAR水平是HIV-1感染患者生存的独立预测因素,对预测疾病的严重程度具有重要参考价值。suPAR在肾脏疾病中也明显上调,被认为是肾脏疾病的可靠生物标志物和疾病进展的致病因素。uPAR在脑缺血或损伤、脓毒症、多发性硬化和风湿性疾病中亦上调。
在衰老过程中也发现uPAR被广泛诱导,suPAR由衰老细胞分泌作为衰老相关分泌表型(SASP)的一部分,诱导衰老的治疗可导致uPAR阳性细胞数量和血清suPAR水平增加,且uPAR阳性细胞表现出与表达衰老相关β半乳糖苷酶(SA-β-gal)的细胞相同的组织学表现,并共表达衰老相关标志物p16和IL-6。在患有衰老相关疾病(如阿尔茨海默病、动脉粥样硬化、骨关节炎、糖尿病和特发性肺纤维化)的患者中,uPAR和/或suPAR的水平也有明显升高。2020年发表在Nature上的研究发现靶向uPAR的CAR T细胞有效逆转了衰老相关病理过程,表明uPAR是潜在的衰老细胞的靶标。
此外,uPAR还作为信号受体发挥作用,通过细胞内信号传导影响迁移、粘附、分化和增殖从而促进肿瘤细胞的运动、侵袭和存活。在前列腺癌、乳腺癌、结直肠癌等几种类型的肿瘤中,uPAR和suPAR水平升高与不良预后密切相关。靶向uPAR是目前肿瘤治疗药物的开发热点,近年来备受关注。
目前研究构建了可靶向uPAR的偶联药物、成像剂和纳米药物递送载体,其靶向性依托uPA-uPAR结合特性设计出可结合uPAR的多肽(如AE105、U11)和抗体(如ATN-658、2G10),虽呈现出一定的靶向诊疗效果,但仍存在许多问题,如多肽的亲和度低、降解代谢快、稳定性差、特异性低;抗体分子量过大而在体内清除缓慢、半衰期长和偶联受阻,且部分抗体因偶联可逆大大降低了抗体的稳定性及配对亲和度等。
因此,设计出能够靶向结合uPAR且亲和度高、稳定性好、分子量小、特异性高的分子,有助于更有效检测血液及组织中uPAR的表达水平、有助于精准对uPAR高表达病灶进行成像或给药,对肿瘤疾病、炎症性疾病、衰老相关疾病等多种疾病的诊断、治疗及科学研究具有重大意义。
发明内容
本发明的目的在于针对现有技术的不足,提供一种识别uPAR的纳米抗体及纳米抗体的相关应用。
本发明所采取的技术方案是:
本发明的第一个方面,提供一种抗uPAR的纳米抗体或其抗原结合片段,所述纳米抗体或其抗原结合片段包含重链;所述重链包含:重链可变区,
所述重链可变区具有:
如SEQ ID NO:2所示的CDR1、如SEQ ID NO:3所示的CDR2和如SEQ ID NO:4所示的CDR3;
或,如SEQ ID NO:6所示的CDR1、如SEQ ID NO:7所示的CDR2和如SEQ ID NO:8所示的CDR3;
或,如SEQ ID NO:10所示的CDR1、如SEQ ID NO:11所示的CDR2和如SEQ ID NO:12所示的CDR3;
或,如SEQ ID NO:14所示的CDR1、如SEQ ID NO:15所示的CDR2和如SEQ ID NO:16所示的CDR3。
更具体地,根据本发明第一个方面所述的纳米抗体或其抗原结合片段,所述重链可变区具有:
如SEQ ID NO:1所示的氨基酸序列;
或,如SEQ ID NO:5所示的氨基酸序列;
或,如SEQ ID NO:9所示的氨基酸序列;
或,如SEQ ID NO:13所示的氨基酸序列。
本发明的第二个方面,提供一种重组蛋白,包含:本发明第一个方面所述的纳米抗体或其抗原结合片段;和
任选的协助表达和/或纯化的标签序列。
本发明的第三个方面,提供与本发明第一个方面所述的纳米抗体或其抗原结合片段、或本发明第二个方面所述的重组蛋白相关的生物材料,所述生物材料包含b1)~b16)中至少一种:
b1)编码本发明第一个方面所述的纳米抗体或其抗原结合片段、或本发明第二个方面所述的重组蛋白的核酸分子;
b2)包含b1)所述核酸分子的表达盒;
b3)包含b1)所述核酸分子的载体;
b4)包含b2)所述表达盒的载体;
b5)包含b1)所述核酸分子的转基因细胞系;
b6)包含b2)所述表达盒的转基因细胞系;
b7)包含b3)所述载体的转基因细胞系;
b8)包含b4)所述载体的转基因细胞系;
b9)包含b1)所述核酸分子的微生物;
b10)包含b2)所述表达盒的微生物;
b11)包含b3)所述载体的微生物;
b12)包含b4)所述载体的微生物;
b13)包含b1)所述核酸分子的病毒;
b14)包含b2)所述表达盒的病毒;
b15)包含b3)所述载体的病毒;
b16)包含b4)所述载体的病毒。
本发明的第四个方面,提供一种偶联物,包含:本发明第一个方面所述的纳米抗体或其抗原结合片段、或本发明第二个方面所述的重组蛋白中的至少一种;
以及偶联部分,所述偶联部分包含可检测标记物、药物、毒素、细胞因子、放射性核素、酶中的至少一种。
本发明的第五个方面,提供(Ⅰ)~(Ⅳ)中至少一项在制备产品中的应用:
(Ⅰ)本发明第一个方面所述的纳米抗体或其抗原结合片段;
(Ⅱ)本发明第二个方面所述的重组蛋白;
(Ⅲ)本发明第三个方面所述的生物材料;
(Ⅳ)本发明第四个方面所述的偶联物;
所述产品包含药物、试剂、检测板、试剂盒、检测芯片中的至少一种。
进一步地,根据本发明第五个方面所述的应用,所述药物具有c1)~c2)中至少一种功能;
c1)靶向治疗肿瘤;
c2)靶向治疗衰老细胞。
uPAR蛋白是膜蛋白,研究表明uPAR蛋白在肿瘤细胞和衰老细胞表面高表达,可作为靶向肿瘤细胞或衰老细胞的药物靶点。
进一步地,根据本发明第五个方面所述的应用,所述试剂、检测板、试剂盒、检测芯片具有d1)~d4)中至少一项功能;
d1)检测uPAR蛋白在样品中的存在或水平;
d2)检测肿瘤细胞;
d3)检测衰老细胞;
d4)诊断疾病预后情况。
更进一步地,所述疾病包括:肿瘤、炎性、感染性疾病、心血管疾病。
本发明个的第六个方面,提供一种产品,所述产品包含h1)~h3)中的至少一种:
h1)本发明第一个方面所述的单克隆抗体或其抗原结合片段;
h2)本发明第二个方面所述的重组蛋白;
h3)本发明第四个方面所述的偶联物;
所述产品包含试剂、检测板、试剂盒、检测芯片中的至少一种。
本发明的第七个方面,提供一种药物,所述药物包含(l1)~(l4)中的至少一种:
(l1)本发明第一个方面所述的单克隆抗体或其抗原结合片段;
(l2)本发明第二个方面所述的重组蛋白;
(l3)本发明第三个方面所述的生物材料;
(l4)本发明第四个方面所述的偶联物;
优选地,所述药物还包含药学上可接受的载体。
本发明第八个方面,提供一种疫苗,所述疫苗包含(l1)~(l4)中至少一种和佐剂:
(l1)本发明第一个方面所述的单克隆抗体或其抗原结合片段;
(l2)本发明第二个方面所述的重组蛋白;
(l3)本发明第三个方面所述的生物材料;
(l4)本发明第四个方面所述的偶联物。
本发明的有益效果是:
本发明提供了一种识别uPAR的纳米抗体或其抗原结合片段,该纳米抗体或其抗原结合片段对uPAR蛋白具有较高的亲和力,相比于传统抗体技术具有特异性强、亲和力高、分子量小、可溶性高、稳定性强、结构简单、基因工程易改造等优势,为多种疾病的诊疗及研究提供了更优良的支持。
可以作为高反应性的检测探针检测血液和组织中的suPAR/uPAR表达,对肿瘤疾病、炎性疾病、感染性疾病、心血管疾病等多种病症的严重程度及预后评判有重要作用,对临床诊疗方案制定具有重要参考价值。近年来的研究还表明uPAR在衰老、部分肿瘤及炎性疾病病理过程中被广泛诱导,其可作为衰老细胞、肿瘤细胞和炎性组织表面标志物,因此uPAR纳米抗体还可作为检测体内衰老细胞、肿瘤细胞及炎性组织的探针,当其偶联成像剂时还可进行定位成像。同理在衰老、肿瘤等靶向药物的设计中,uPAR纳米抗体亦可作为靶向药物的眼睛和抓手,能够准确识别并结合uPAR从而定位衰老细胞和肿瘤细胞进行靶向作用或给药。综上,本发明对肿瘤疾病、炎性疾病、感染性疾病、心血管疾病等多种疾病的检测、诊断、治疗及研究都具有重要意义。
附图说明
图1 四轮噬菌体筛选过程中检测每轮筛选后富集到的纳米抗体库容量,富集到的噬菌体梯度稀释后感染TG1菌株接着在氨苄平板下培养后计数计算库容量。
图2第四轮富集后的纳米抗体文库中挑取96个进行表达并ELISA检测其与uPAR的亲和力。注:检测结果按ELISA信号强弱从左到右排列,由于克隆数量太多,图中并未显示每个克隆的编号。
具体实施方式
以下通过具体的实施例对本发明的内容作进一步详细的说明。
应理解,这些实施例仅用于说明本发明而不用于限制本发明的范围。
下列实施例中未注明具体条件的实验方法,通常按照常规条件,或按照制造厂商所建议的条件。本实施例中所使用的材料、试剂等,如无特别说明,为从商业途径得到的试剂和材料。
实施例1uPAR纳米抗体的筛选/制备
发明人团队用纯化的uPAR作为抗原对已经构建好的纳米抗体文库进行筛选,每轮筛选均使用1×1012噬菌体,每轮筛选到的噬菌体检测滴度后进行扩增培养用于下一轮筛选。
从第三轮开始获得的噬菌体显著增加,表明随着筛选的进行对uPAR抗原具有较高亲和力的噬菌体被逐步富集,在最后一轮筛选中获得3.2×109噬菌体。随机挑取96个纳米抗体进行表达,并通过ELISA验证纳米抗体对uPAR的识别能力,结果表明大于80%的抗体对uPAR抗原呈阳性反应,其中第四轮富集的纳米抗体文库中80%以上是uPAR特异性的纳米抗体。
TG1文库的构建:从新鲜羊驼血液中提取分离PBMC;用Trizol法提取RNA;将提取出来的RNA全部反转录成cDNA,以cDNA为模板扩增进行2轮巢式PCR;SfiI&NotI酶切巢式pcr第二轮产物后与酶切的载体进行连接,过夜后的连接产物进行超滤浓缩纯化;纯化后的连接产物加入事先做好的TG1电转感受态进行电转,37℃培养1h,复苏液进行梯度稀释测电转库容,并将所有复苏液涂于适量大平板中37℃培养过夜;大平板上的菌全部刮下收集起来加甘油-80℃保存备。VHH噬菌体文库的扩增:20 OD的TG1文库加入200 ml 2YT(A&G)培养基,37℃ 220 rpm/min扩增,至OD值为0.4~0.6;加入20倍数量辅助噬菌体M13K07摇匀,37℃静置30 min进行侵染;接着37℃ 220 rpm/min 1h;5000 g 离心 5 min,弃旧培养基;离心后的沉淀用等体积2YT(A&K)重悬 30℃ 220 rpm/min过夜让噬菌体扩增;过夜后的培养液用5000 g 离心10 min;留上清,上清中加1/5体积PEG-NaCl,冰上静置2 h以上以让噬菌体形成沉淀;8000 g 4℃离心10 min,弃上清,留沉淀(含噬菌体);沉淀用5 mL PBS溶解,8000 g4 ℃离心10min留上清,弃沉淀;上清中加1/5体积PEG-NaCl,4℃静置1 h以上再次沉淀噬菌体;8000g 4℃离心10min;弃上清,留沉淀,沉淀用适量PBS溶解;测滴度待用。
生物淘洗:星形管包被人uPAR (50 μg/管) 4℃,5 % MPBS封闭,37℃静置2h;加入VHH噬菌体(1×10^12,5%MPBS中37℃旋转台上孵浴0.5 h,静置1.5 h);用0.05 % PBST清洗10次,用PBS清洗5次。洗脱,中和,侵染,测滴度。
扩增第一轮淘洗的噬菌体:接种TG1到10 mL 2YT-A&G培养基中,37℃ 220 rpm/min培养至OD值为0.4~0.6;加入洗脱液(600ul)混匀,37 ℃静置30 min,37℃ 220 rpm/min30min;加入30 mL 2YT-A&G培养基,37℃ 220 rpm/min培养至OD值为0.4~0.6;加入20倍过量的M13K07辅助噬菌体,37℃静置30 min,37℃ 220 rpm/min培养30~60 min;5000 rpm5min离心,弃旧培养基;离心后的沉淀用等体积2YT(Amp+&Kan+)重悬,30℃ 220 rpm/min过夜;过夜后的培养液5000 g离心10 min留上清,上清中加1/5体积PEG-NaCl,冰上静置2h以上沉淀噬菌体;8000 g 4℃离心10 min弃上清,留沉淀,沉淀用5mL PBS溶解;8000 g 4℃离心10 min留上清,弃沉淀;上清中再次加1/5体积PEG-NaCl,4℃静置1h以上沉淀噬菌体;8000g,4℃离心10 min,弃上清,留沉淀,沉淀用适量PBS溶解;测定滴度。
进行第二到第四轮生物淘洗和淘洗后噬菌体的扩增,每轮淘洗投入的噬菌体均为1×10^12次方。每轮淘洗后获得的噬菌体数目见下表1及附图1:
表1四轮噬菌体筛选过程中噬菌体文库数量
注:每轮筛选时投入1×1012的噬菌体(Input)与uPAR抗原包被的ELISA板子结合,随后统计洗脱的噬菌体(Output)并计算回收率。
实施例2 纳米抗体的鉴定
为了进一步验证哪些纳米抗体具有识别uPAR的能力,发明人团队从第4轮富集测滴度的平板中挑96个单克隆接种到2YT-A&G液体培养基中,37℃ 220 rpm/min培养至OD值为0.4~0.6,取100uL保种,加入20倍数量的M13K07辅助噬菌体,37℃静置30min,37℃ 220rpm/min 30~60 min 3800rpm 5min离心,弃旧培养基离心后的沉淀用等体积2YT(Amp+&Kan+)重悬 30℃ 220 rpm/min 过夜;过夜后的培养液3800 rpm 离心10min,留上清,上清中包含噬菌体并用于后续Elisa检测噬菌体与uPAR的相互作用。
ELISA检测纳米抗体噬菌粒与uPAR的相互作用:
uPAR蛋白按100 ng/孔包被ELISA板,37 ℃静置120 min,弃包被液,PBS清洗3次;4% MPBS封闭,4 ℃过夜,弃封闭液,PBS清洗3次;8 % MPBS 50 μL+离心后的上清培养基50 μL混匀,37 ℃旋转台上孵育0.5 h,37 ℃静置1 h,弃培养液,0.05 % PBST清洗3次+PBS清洗3次;4 % MPBS稀释M13-Anti-HRP,100 μL/孔,37℃静置1h,弃M13-Anti-HRP,0.05 % PBST清洗3次+PBS清洗3次;加入TMB系统显色液100 μL/孔,37℃静置5 min,1M HCl终止。结果表明大于80%的抗体对uPAR抗原呈阳性反应(见附图2所示),表明第四轮富集的纳米抗体文库中80%以上是uPAR特异性的纳米抗体。进一步将ELISA阳性信号排前20的克隆进行sanger测序,其中部分序列是相同的,最终得到4个uPAR特异性纳米抗体的氨基酸序列(下划线对应重链可变区中的高变区序列,依次为CDR1、CDR2、CDR3)。
其中,>3B1的氨基酸序列如下所示:
QLQLVESGGGLVQPGGSLRLSCAASGSIFSINTMGWHRQAPGKQREFVAGITSAGSANYADSVKGRFTISRDNAKNTVYLQMNSLKPEDTAVYYCNADWGIIMRNWGQGTQVTVSSAHHSEDPAAA(SEQ ID NO:1)。
>3A11的氨基酸序列如下所示:
QLQLVESGGGSVQAGGSLRLSCAASGSMFGIKIMGWFRQAPGKEREFVAGVSRFAGNTLYADSVKGRFTVSRDNARSTVYLQMNSLKPDDTAIYYCAAGRWVAVTTTPVRYEYWGQGTQVTVSSAHHSEDPAAA(SEQ ID NO:5)。
>3A8的氨基酸序列如下所示:
QVQLVESGGGLVQPGGSLRLSCAASGFTFSSYYMSWVRQAPGKGPEWVSRINAGGSRTSYADSVQGRFTISRDNAKNTLYLQMNSLKPEDTAVYYCNAHIGTSSYARTVYWGQGTQVTVSSAHHSEDPAAA(SEQ ID NO:9)。
>3A6的氨基酸序列如下所示:
QVQLVESGGGLVQAGGSLTLSCKASGFEFSNYNLGWFRQAPGKEREFVASFGWSADYADSAKGRFTISRDRRKNTLYLQMNNLKPEDTAVYYCAGVFLGVGSTFRRARDAQYWGQGTQVTVSSAHHSEDPAAA(SEQ ID NO:13)。
综上,发明人团队提供了可识别uPAR的纳米抗体或其抗原结合片段,该纳米抗体或其抗原结合片段对uPAR蛋白具有较高的亲和力,相比于传统抗体技术具有特异性强、亲和力高、分子量小、可溶性高、稳定性强、结构简单、基因工程易改造等优势,可以作为高反应性的检测探针检测血液和组织中的suPAR/uPAR表达,对肿瘤疾病、炎性疾病、感染性疾病、心血管疾病等多种病症的严重程度及预后评判有重要作用,对临床诊疗方案制定具有重要参考价值。近年来的研究还表明uPAR在衰老、部分肿瘤及炎性疾病病理过程中被广泛诱导,其可作为衰老细胞、肿瘤细胞和炎性组织表面标志物,因此uPAR纳米抗体还可作为检测体内衰老细胞、肿瘤细胞及炎性组织的探针,当其偶联成像剂时还可进行定位成像。同理在衰老、肿瘤等靶向药物的设计中,uPAR纳米抗体亦可作为靶向药物的眼睛和抓手,能够准确识别并结合uPAR从而定位衰老细胞和肿瘤细胞进行靶向作用或给药。
上述实施例为本发明较佳的实施方式,但本发明的实施方式并不受上述实施例的限制,其他的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化,均应为等效的置换方式,都包含在本发明的保护范围之内。
Claims (10)
1.一种抗uPAR的纳米抗体或其抗原结合片段,所述纳米抗体或其抗原结合片段包含重链;所述重链包含:重链可变区,
所述重链可变区具有:
如SEQ ID NO:2所示的CDR1、如SEQ ID NO:3所示的CDR2和如SEQ ID NO:4所示的CDR3;
或,如SEQ ID NO:6所示的CDR1、如SEQ ID NO:7所示的CDR2和如SEQ ID NO:8所示的CDR3;
或,如SEQ ID NO:10所示的CDR1、如SEQ ID NO:11所示的CDR2和如SEQ ID NO:12所示的CDR3;
或,如SEQ ID NO:14所示的CDR1、如SEQ ID NO:15所示的CDR2和如SEQ ID NO:16所示的CDR3。
2.根据权利要求1所述的纳米抗体或其抗原结合片段,其特征在于,所述重链可变区具有:
如SEQ ID NO:1所示的氨基酸序列;
或,如SEQ ID NO:5所示的氨基酸序列;
或,如SEQ ID NO:9所示的氨基酸序列;
或,如SEQ ID NO:13所示的氨基酸序列。
3.一种重组蛋白,包含:权利要求1或2所述的纳米抗体或其抗原结合片段;和
协助表达和/或纯化的标签序列。
4.与权利要求1或2所述的纳米抗体或其抗原结合片段、或权利要求3所述的重组蛋白相关的生物材料,所述生物材料为b1)~b16)中至少一种:
b1)编码权利要求1或2所述的纳米抗体或其抗原结合片段、或权利要求3所述的重组蛋白的核酸分子;
b2)包含b1)所述核酸分子的表达盒;
b3)包含b1)所述核酸分子的载体;
b4)包含b2)所述表达盒的载体;
b5)包含b1)所述核酸分子的转基因细胞系;
b6)包含b2)所述表达盒的转基因细胞系;
b7)包含b3)所述载体的转基因细胞系;
b8)包含b4)所述载体的转基因细胞系;
b9)包含b1)所述核酸分子的微生物;
b10)包含b2)所述表达盒的微生物;
b11)包含b3)所述载体的微生物;
b12)包含b4)所述载体的微生物;
b13)包含b1)所述核酸分子的病毒;
b14)包含b2)所述表达盒的病毒;
b15)包含b3)所述载体的病毒;
b16)包含b4)所述载体的病毒。
5.一种偶联物,包含:权利要求1或2所述的纳米抗体或其抗原结合片段、或权利要求3所述的重组蛋白中的至少一种;
以及偶联部分,所述偶联部分选自可检测标记物、药物、毒素、细胞因子、放射性核素、酶中的至少一种。
6.(Ⅰ)~(Ⅳ)中至少一项在制备产品中的应用:
(Ⅰ)权利要求1或2所述的纳米抗体或其抗原结合片段;
(Ⅱ)权利要求3所述的重组蛋白;
(Ⅲ)权利要求4所述的生物材料;
(Ⅳ)权利要求5所述的偶联物;
所述产品包含药物、试剂、检测板、试剂盒、检测芯片中的至少一种;
所述产品具有检测uPAR蛋白在样品中的存在或水平的功能。
7.根据权利要求6所述的应用,其特征在于,所述药物具有c1)~c2)中至少一种功能;
c1)靶向肿瘤细胞;
c2)靶向衰老细胞。
8.根据权利要求6所述的应用,其特征在于,所述试剂、检测板、试剂盒、检测芯片还具有d1)~d3)中至少一项功能;
d1)检测肿瘤细胞;
d2)检测衰老细胞;
d3)诊断疾病预后情况。
9.一种产品,所述产品包含h1)~h3)中的至少一种:
h1)权利要求1或2所述的纳米抗体或其抗原结合片段;
h2)权利要求3所述的重组蛋白;
h3)权利要求5所述的偶联物;
所述产品包含试剂、检测板、试剂盒、检测芯片中的至少一种。
10.一种药物,所述药物包含l1)~l4)中的至少一种:
l1)权利要求1或2所述的纳米抗体或其抗原结合片段;
l2)权利要求3所述的重组蛋白;
l3)权利要求4所述的生物材料;
l4)权利要求5所述的偶联物。
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