CN110437326B - 大疱性类天疱疮BP230 IgE抗体的抗原表位多肽及其用途 - Google Patents

大疱性类天疱疮BP230 IgE抗体的抗原表位多肽及其用途 Download PDF

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CN110437326B
CN110437326B CN201910755415.2A CN201910755415A CN110437326B CN 110437326 B CN110437326 B CN 110437326B CN 201910755415 A CN201910755415 A CN 201910755415A CN 110437326 B CN110437326 B CN 110437326B
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元慧杰
潘萌
史晏绮
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Ruinjin Hospital Affiliated to Shanghai Jiaotong University School of Medicine Co Ltd
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Abstract

本发明涉及一种大疱性类天疱疮BP230 IgE型抗体的抗原表位多肽及其用途，具体而言，本发明提供了大疱性类天疱疮BP230 IgE型抗体的抗原表位多肽，本发明还提供了可特异性结合上述表位多肽的结合分子、包含所述结合分子的抗血清、检测大疱性类天疱疮的试剂盒、检测方法及抑制大疱性类天疱疮的组合物；本发明还提供了上述表位多肽、结合分子和抗血清在制备预防、治疗大疱性类天疱疮疾病的药物以及大疱性类天疱疮诊断剂中的用途。其优点表现在：本发明首次定位特异性IgE型抗体与BP230抗原片段结合的表位。这对于大疱性类天疱疮疫苗及诊断试剂盒的开发有重要的指导意义。

Description

大疱性类天疱疮BP230 IgE抗体的抗原表位多肽及其用途
技术领域
本发明涉及生物技术领域，具体地说，是大疱性类天疱疮BP230 IgE抗体的抗原表位多肽及其用途。
背景技术
大疱性类天疱疮(BP)是一种常见的自身免疫性大疱病。其特征为患者血清中存在特异性抗BP180和BP230抗体。这些抗体与基底膜带BP抗原结合，通过激活补体，吸引炎性细胞并释放蛋白水解酶，导致表皮与基底膜带粘附能力丧失而致水疱形成。
在以往大疱性类天疱疮的研究中，主要是检测特异性抗BP180和BP230的IgG型抗体，其中BP230 IgG型抗体在BP230上的结合位点为C段(氨基酸序列为1869-2649)，其次是N段(氨基酸序列为1-979)，目前包被这两段抗原片段的ELISA检测试剂盒为MBL公司生产，另也有欧蒙公司生产的单独包被C段的ELISA试剂盒。而越来越多研究表明大疱性类天疱疮患者血清中有特异性抗BP180和BP230的IgE型抗体，目前大部分的特异性抗BP230 IgE型抗体的检测是直接沿用MBL公司生产的ELISA试剂盒，仅包含BP230为N段和C段，或者部分研究只检测BP230的部分片段(主要是C段)，并不是BP230抗原的全长，所以不能很好的确立病人血清中IgE型抗体在BP230中的主要结合位点，也造成在检测特异性抗BP230 IgE型抗体在患者中检出的阳性率降低。
截止目前为止，大疱性类天疱疮BP230 IgE抗体的抗原表位仍未被鉴定出，而大疱性类天疱疮BP230 IgE抗体的抗原表位的鉴定对于大疱性类天疱疮的预防和治疗药物以及诊断试剂的制备，其意义十分重大。
因此，发明人设计了本发明大疱性类天疱疮BP230 IgE抗体的抗原表位多肽及其用途，关于此内容目前还未见报道。
发明内容
本发明的第一个目的是针对现有技术的不足，提供一种分离的多肽。
本发明的第二个目的是针对现有技术的不足，提供上述多肽的用途。
本发明的第三个目的是针对现有技术的不足，提供一种基于上述多肽的结合分子。
本发明的第四个目的是针对现有技术的不足，提供上述结合分子的用途。
本发明的第五个目的是针对现有技术的不足，提供一种抗血清。
本发明的第六个目的是针对现有技术的不足，提供上述抗血清的用途。
本发明的第七个目的是针对现有技术的不足，提供一种检测大疱性类天疱疮的试剂盒。
本发明的第八个目的是针对现有技术的不足，提供一种非治疗目的的检测大疱性类天疱疮BP230 IgE型抗体的方法。
本发明的第九个目的是针对现有技术的不足，提供一种抑制大疱性类天疱疮的组合物。
为实现上述第一个目的，本发明采取的技术方案是：
一种分离的多肽，其氨基酸序列如SEQ ID NO.2所示。
为实现上述第二个目的，本发明采取的技术方案是：
如上所述的多肽在制备预防或治疗大疱性类天疱疮疾病的药物，或制备大疱性类天疱疮疾病诊断剂中的用途。
为实现上述第三个目的，本发明采取的技术方案是：
一种结合分子，所述的结合分子可特异性结合如上所述的多肽。
作为本发明的一个优选实施方案，所述的结合分子是抗体。
为实现上述第四个目的，本发明采取的技术方案是：
如上所述的结合分子在制备预防或治疗大疱性类天疱疮疾病的药物，或制备大疱性类天疱疮疾病诊断剂中的用途。
为实现上述第五个目的，本发明采取的技术方案是：
一种抗血清，所述的抗血清包含如上所述的结合分子。
为实现上述第六个目的，本发明采取的技术方案是：
如上的抗血清在制备预防或治疗大疱性类天疱疮疾病的药物，或制备大疱性类天疱疮疾病诊断剂中的用途。
为实现上述第七个目的，本发明采取的技术方案是：
一种检测大疱性类天疱疮的试剂盒，所述的试剂盒包含如上所述的多肽或结合分子。
为实现上述第八个目的，本发明采取的技术方案是：
一种非治疗目的的检测大疱性类天疱疮BP230 IgE型抗体的方法，包括步骤：
a)将样品与如上所述的多肽接触；
b)检测是否形成抗原-抗体复合物，其中形成复合物就表示样品中存在大疱性类天疱疮BP230 IgE型抗体。
为实现上述第九个目的，本发明采取的技术方案是：
一种抑制大疱性类天疱疮的组合物，所述的组合物包含：
a)如上所述的结合分子；和
b)药学上可接受的载体。
本发明优点在于：
通过检测大疱性类天疱疮患者血清中特异性抗7段重组BP230抗原片段的IgE型抗体，结合相关统计学分析，定位特异性IgE型抗体在BP230抗原片段的表位。提高抗BP230IgE型抗体在大疱性类天疱疮患者中检出的阳性率，为大疱性类天疱疮疾病的诊疗提供理论依据和新靶点。
附图说明
附图1是BP230-R1段与其他段抗原蛋白IgE型抗体水平的比较。
具体实施方式
下面结合具体实施方式，进一步阐述本发明。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。此外应理解，在阅读了本发明记载的内容之后，本领域技术人员可以对本发明作各种改动或修改，这些等价形式同样落于本申请所附权利要求书所限定的范围。
BP230抗原：DystoninProteinLength＝2649MW＝306702.6pI＝6.25
其氨基酸序列的氨基酸序列如SEQ ID NO.1所示。
R1氨基酸序列的氨基酸序列如SEQ ID NO.2所示。
实施例1表位多肽的合成、筛选和鉴定
一、实验材料：
1.重组人BP230的7段抗原片段(德泰生物科技)：将BP230分为7段，分别为：N1(氨基酸序列为2-443)，N2(氨基酸序列为432-864)，R1(氨基酸序列为846-1297)，R2(氨基酸序列为1286-1742)，C1(氨基酸序列为1710-2033)，C2(氨基酸序列为2024-2342)，C3(氨基酸序列为2326-2649)。均为融合C-His标签，来源于哺乳动物表达系统，瞬时转染表达细胞为HEK293，通过亲和层析纯化获得，蛋白储存液为PBS缓冲体系(含5％甘油)，pH＝7.4，-80℃冰箱保存。
2.抗原包被液：1L蒸馏水内含1.59克Na2CO3，2.93克NaHCO3和0.2克NaN3，pH＝9.6，4℃冰箱保存。
3.96孔板(CorningIncorporated)
4.患者血清：42名大疱性类天疱疮患者和10名正常人血清。
5.抗原封闭液/血清稀释液：1×TBS溶液内含2％胎牛血清和0.01％Tween20，4℃冰箱保存。
6.HRP标记的鼠抗人IgE(Southernbiotec)
7.清洗缓冲液(MBL)：含PBS和Tween20的10倍浓缩液，4℃冰箱保存。
8.酶基质液(MBL)：TMB/H2O2溶液，4℃冰箱保存。
9.终止液(MBL)：1.0N硫酸溶液，4℃冰箱保存。
二、实验仪器：
酶标仪(波长：450nm)
三、实验方法：
1.BP230的7段抗原片段包被于96孔板：将BP230的7段片段分别用抗原包被液稀释至浓度4ug/mL，每孔100ul加入96孔板内，存放于4℃冰箱内过夜。
2.封闭抗原：将96孔板内的抗原蛋白弃去后，用1×清洗缓冲液洗涤4次后，每孔加入抗原封闭液200ul，室温封闭2小时后，弃去封闭液，洗涤4次。
3.ELISA方法检测患者的7段BP230抗原IgE型抗体(表位定位)：血清以1:20浓度稀释，每孔100ul加入96孔板，于37度水浴孵育60分钟后，洗涤4次；加入二抗，二抗为HRP标记的鼠抗人IgE，稀释浓度为1:1000，每孔100ul加入96孔板，于37℃水浴孵育45分钟后，洗涤4次；加入酶基质物，每孔100ul加入96孔板，于室温孵育20分钟后，每孔100ul加入终止液。放于450nm酶标仪中进行读数。
4.统计学方法：Cut-off值为10名正常人测定的OD值的平均数+3个标准差(mean+3SD)。计算7段BP230抗原蛋白的阳性率。通过Graph Pad Prism5软件，运用Mann-Whitney方法，对BP230各段IgE型抗体水平进行差异性分析。
四、实验结果
1、病人以抗BP230-R1 IgE型抗体为主：用ELISA方法检测，根据Cut-off值界定各段IgE型抗体的阳性OD值，各段IgE型抗体的阳性率分别为0.33％(N1)，0.38％(N2)，0.55％(R1)，0.38％(R2)0.33％(C1)，0.31％(C2)，0.29％(C3)，结果发现患者中以抗BP230-R1IgE阳性率最高(见表1)。
表1、7段BP230抗原蛋白IgE型抗体的阳性率
Figure BDA0002168564280000051
2、抗BP230-R1 IgE型抗体水平较其他段显著升高：将BP230-R1段与其他6段抗原蛋白IgE型抗体水平进行差异性分析，结果发现除了BP230-R1与BP230-N2 IgE型抗体水平之间无显著性差异之外，其余BP230-N1、BP230-R2、BP230-C1、BP230-C2、BP230-C3的IgE型抗体水平均有显著性差异，以BP230-R1 IgE型抗体水平较高(见图1)。
根据以上两个实验结果，我们认为抗BP230的IgE型抗体主要是与BP230的R1段表位结合。
本发明通过检测大疱性类天疱疮患者血清中特异性抗7段重组BP230抗原片段的IgE型抗体，结合相关统计学分析，定位特异性IgE型抗体在BP230抗原片段的表位。提高抗BP230 IgE型抗体在大疱性类天疱疮患者中检出的阳性率，为此病的诊疗提供理论依据和新靶点。
以上所述仅是本发明的优选实施方式，应当指出，对于本技术领域的普通技术人员，在不脱离本发明原理的前提下，还可以做出若干改进和补充，这些改进和补充也应视为本发明的保护范围。
SEQUENCE LISTING
<110> 上海交通大学医学院附属瑞金医院
<120> 大疱性类天疱疮BP230 IgE抗体的抗原表位多肽及其用途
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930 935 940
Trp Gln Asn Val His Val Gln Ile Asp Asn Arg Leu Arg Asp Leu Glu
945 950 955 960
Gly Ile Gly Lys Ser Leu Lys Tyr Tyr Arg Asp Thr Tyr His Pro Leu
965 970 975
Asp Asp Trp Ile Gln Gln Val Glu Thr Thr Gln Arg Lys Ile Gln Glu
980 985 990
Asn Gln Pro Glu Asn Ser Lys Thr Leu Ala Thr Gln Leu Asn Gln Gln
995 1000 1005
Lys Met Leu Val Ser Glu Ile Glu Met Lys Gln Ser Lys Met Asp
1010 1015 1020
Glu Cys Gln Lys Tyr Ala Glu Gln Tyr Ser Ala Thr Val Lys Asp
1025 1030 1035
Tyr Glu Leu Gln Thr Met Thr Tyr Arg Ala Met Val Asp Ser Gln
1040 1045 1050
Gln Lys Ser Pro Val Lys Arg Arg Arg Met Gln Ser Ser Ala Asp
1055 1060 1065
Leu Ile Ile Gln Glu Phe Met Asp Leu Arg Thr Arg Tyr Thr Ala
1070 1075 1080
Leu Val Thr Leu Met Thr Gln Tyr Ile Lys Phe Ala Gly Asp Ser
1085 1090 1095
Leu Lys Arg Leu Glu Glu Glu Glu Ile Lys Arg Cys Lys Glu Thr
1100 1105 1110
Ser Glu His Gly Ala Tyr Ser Asp Leu Leu Gln Arg Gln Lys Ala
1115 1120 1125
Thr Val Leu Glu Asn Ser Lys Leu Thr Gly Lys Ile Ser Glu Leu
1130 1135 1140
Glu Arg Met Val Ala Glu Leu Lys Lys Gln Lys Ser Arg Val Glu
1145 1150 1155
Glu Glu Leu Pro Lys Val Arg Glu Ala Ala Glu Asn Glu Leu Arg
1160 1165 1170
Lys Gln Gln Arg Asn Val Glu Asp Ile Ser Leu Gln Lys Ile Arg
1175 1180 1185
Ala Glu Ser Glu Ala Lys Gln Tyr Arg Arg Glu Leu Glu Thr Ile
1190 1195 1200
Val Arg Glu Lys Glu Ala Ala Glu Arg Glu Leu Glu Arg Val Arg
1205 1210 1215
Gln Leu Thr Ile Glu Ala Glu Ala Lys Arg Ala Ala Val Glu Glu
1220 1225 1230
Asn Leu Leu Asn Phe Arg Asn Gln Leu Glu Glu Asn Thr Phe Thr
1235 1240 1245
Arg Arg Thr Leu Glu Asp His Leu Lys Arg Lys Asp Leu Ser Leu
1250 1255 1260
Asn Asp Leu Glu Gln Gln Lys Asn Lys Leu Met Glu Glu Leu Arg
1265 1270 1275
Arg Lys Arg Asp Asn Glu Glu Glu Leu Leu Lys Leu Ile Lys Gln
1280 1285 1290
Met Glu Lys Asp Leu Ala Phe Gln Lys Gln Val Ala Glu Lys Gln
1295 1300 1305
Leu Lys Glu Lys Gln Lys Ile Glu Leu Glu Ala Arg Arg Lys Ile
1310 1315 1320
Thr Glu Ile Gln Tyr Thr Cys Arg Glu Asn Ala Leu Pro Val Cys
1325 1330 1335
Pro Ile Thr Gln Ala Thr Ser Cys Arg Ala Val Thr Gly Leu Gln
1340 1345 1350
Gln Glu His Asp Lys Gln Lys Ala Glu Glu Leu Lys Gln Gln Val
1355 1360 1365
Asp Glu Leu Thr Ala Ala Asn Arg Lys Ala Glu Gln Asp Met Arg
1370 1375 1380
Glu Leu Thr Tyr Glu Leu Asn Ala Leu Gln Leu Glu Lys Thr Ser
1385 1390 1395
Ser Glu Glu Lys Ala Arg Leu Leu Lys Asp Lys Leu Asp Glu Thr
1400 1405 1410
Asn Asn Thr Leu Arg Cys Leu Lys Leu Glu Leu Glu Arg Lys Asp
1415 1420 1425
Gln Ala Glu Lys Gly Tyr Ser Gln Gln Leu Arg Glu Leu Gly Arg
1430 1435 1440
Gln Leu Asn Gln Thr Thr Gly Lys Ala Glu Glu Ala Met Gln Glu
1445 1450 1455
Ala Ser Asp Leu Lys Lys Ile Lys Arg Asn Tyr Gln Leu Glu Leu
1460 1465 1470
Glu Ser Leu Asn His Glu Lys Gly Lys Leu Gln Arg Glu Val Asp
1475 1480 1485
Arg Ile Thr Arg Ala His Ala Val Ala Glu Lys Asn Ile Gln His
1490 1495 1500
Leu Asn Ser Gln Ile His Ser Phe Arg Asp Glu Lys Glu Leu Glu
1505 1510 1515
Arg Leu Gln Ile Cys Gln Arg Lys Ser Asp His Leu Lys Glu Gln
1520 1525 1530
Phe Glu Lys Ser His Glu Gln Leu Leu Gln Asn Ile Lys Ala Glu
1535 1540 1545
Lys Glu Asn Asn Asp Lys Ile Gln Arg Leu Asn Glu Glu Leu Glu
1550 1555 1560
Lys Ser Asn Glu Cys Ala Glu Met Leu Lys Gln Lys Val Glu Glu
1565 1570 1575
Leu Thr Arg Gln Asn Asn Glu Thr Lys Leu Met Met Gln Arg Ile
1580 1585 1590
Gln Ala Glu Ser Glu Asn Ile Val Leu Glu Lys Gln Thr Ile Gln
1595 1600 1605
Gln Arg Cys Glu Ala Leu Lys Ile Gln Ala Asp Gly Phe Lys Asp
1610 1615 1620
Gln Leu Arg Ser Thr Asn Glu His Leu His Lys Gln Thr Lys Thr
1625 1630 1635
Glu Gln Asp Phe Gln Arg Lys Ile Lys Cys Leu Glu Glu Asp Leu
1640 1645 1650
Ala Lys Ser Gln Asn Leu Val Ser Glu Phe Lys Gln Lys Cys Asp
1655 1660 1665
Gln Gln Asn Ile Ile Ile Gln Asn Thr Lys Lys Glu Val Arg Asn
1670 1675 1680
Leu Asn Ala Glu Leu Asn Ala Ser Lys Glu Glu Lys Arg Arg Gly
1685 1690 1695
Glu Gln Lys Val Gln Leu Gln Gln Ala Gln Val Gln Glu Leu Asn
1700 1705 1710
Asn Arg Leu Lys Lys Val Gln Asp Glu Leu His Leu Lys Thr Ile
1715 1720 1725
Glu Glu Gln Met Thr His Arg Lys Met Val Leu Phe Gln Glu Glu
1730 1735 1740
Ser Gly Lys Phe Lys Gln Ser Ala Glu Glu Phe Arg Lys Lys Met
1745 1750 1755
Glu Lys Leu Met Glu Ser Lys Val Ile Thr Glu Asn Asp Ile Ser
1760 1765 1770
Gly Ile Arg Leu Asp Phe Val Ser Leu Gln Gln Glu Asn Ser Arg
1775 1780 1785
Ala Gln Glu Asn Ala Lys Leu Cys Glu Thr Asn Ile Lys Glu Leu
1790 1795 1800
Glu Arg Gln Leu Gln Gln Tyr Arg Glu Gln Met Gln Gln Gly Gln
1805 1810 1815
His Met Glu Ala Asn His Tyr Gln Lys Cys Gln Lys Leu Glu Asp
1820 1825 1830
Glu Leu Ile Ala Gln Lys Arg Glu Val Glu Asn Leu Lys Gln Lys
1835 1840 1845
Met Asp Gln Gln Ile Lys Glu His Glu His Gln Leu Val Leu Leu
1850 1855 1860
Gln Cys Glu Ile Gln Lys Lys Ser Thr Ala Lys Asp Cys Thr Phe
1865 1870 1875
Lys Pro Asp Phe Glu Met Thr Val Lys Glu Cys Gln His Ser Gly
1880 1885 1890
Glu Leu Ser Ser Arg Asn Thr Gly His Leu His Pro Thr Pro Arg
1895 1900 1905
Ser Pro Leu Leu Arg Trp Thr Gln Glu Pro Gln Pro Leu Glu Glu
1910 1915 1920
Lys Trp Gln His Arg Val Val Glu Gln Ile Pro Lys Glu Val Gln
1925 1930 1935
Phe Gln Pro Pro Gly Ala Pro Leu Glu Lys Glu Lys Ser Gln Gln
1940 1945 1950
Cys Tyr Ser Glu Tyr Phe Ser Gln Thr Ser Thr Glu Leu Gln Ile
1955 1960 1965
Thr Phe Asp Glu Thr Asn Pro Ile Thr Arg Leu Ser Glu Ile Glu
1970 1975 1980
Lys Ile Arg Asp Gln Ala Leu Asn Asn Ser Arg Pro Pro Val Arg
1985 1990 1995
Tyr Gln Asp Asn Ala Cys Glu Met Glu Leu Val Lys Val Leu Thr
2000 2005 2010
Pro Leu Glu Ile Ala Lys Asn Lys Gln Tyr Asp Met His Thr Glu
2015 2020 2025
Val Thr Thr Leu Lys Gln Glu Lys Asn Pro Val Pro Ser Ala Glu
2030 2035 2040
Glu Trp Met Leu Glu Gly Cys Arg Ala Ser Gly Gly Leu Lys Lys
2045 2050 2055
Gly Asp Phe Leu Lys Lys Gly Leu Glu Pro Glu Thr Phe Gln Asn
2060 2065 2070
Phe Asp Gly Asp His Ala Cys Ser Val Arg Asp Asp Glu Phe Lys
2075 2080 2085
Phe Gln Gly Leu Arg His Thr Val Thr Ala Arg Gln Leu Val Glu
2090 2095 2100
Ala Lys Leu Leu Asp Met Arg Thr Ile Glu Gln Leu Arg Leu Gly
2105 2110 2115
Leu Lys Thr Val Glu Glu Val Gln Lys Thr Leu Asn Lys Phe Leu
2120 2125 2130
Thr Lys Ala Thr Ser Ile Ala Gly Leu Tyr Leu Glu Ser Thr Lys
2135 2140 2145
Glu Lys Ile Ser Phe Ala Ser Ala Ala Glu Arg Ile Ile Ile Asp
2150 2155 2160
Lys Met Val Ala Leu Ala Phe Leu Glu Ala Gln Ala Ala Thr Gly
2165 2170 2175
Phe Ile Ile Asp Pro Ile Ser Gly Gln Thr Tyr Ser Val Glu Asp
2180 2185 2190
Ala Val Leu Lys Gly Val Val Asp Pro Glu Phe Arg Ile Arg Leu
2195 2200 2205
Leu Glu Ala Glu Lys Ala Ala Val Gly Tyr Ser Tyr Ser Ser Lys
2210 2215 2220
Thr Leu Ser Val Phe Gln Ala Met Glu Asn Arg Met Leu Asp Arg
2225 2230 2235
Gln Lys Gly Lys His Ile Leu Glu Ala Gln Ile Ala Ser Gly Gly
2240 2245 2250
Val Ile Asp Pro Val Arg Gly Ile Arg Val Pro Pro Glu Ile Ala
2255 2260 2265
Leu Gln Gln Gly Leu Leu Asn Asn Ala Ile Leu Gln Phe Leu His
2270 2275 2280
Glu Pro Ser Ser Asn Thr Arg Val Phe Pro Asn Pro Asn Asn Lys
2285 2290 2295
Gln Ala Leu Tyr Tyr Ser Glu Leu Leu Arg Met Cys Val Phe Asp
2300 2305 2310
Val Glu Ser Gln Cys Phe Leu Phe Pro Phe Gly Glu Arg Asn Ile
2315 2320 2325
Ser Asn Leu Asn Val Lys Lys Thr His Arg Ile Ser Val Val Asp
2330 2335 2340
Thr Lys Thr Gly Ser Glu Leu Thr Val Tyr Glu Ala Phe Gln Arg
2345 2350 2355
Asn Leu Ile Glu Lys Ser Ile Tyr Leu Glu Leu Ser Gly Gln Gln
2360 2365 2370
Tyr Gln Trp Lys Glu Ala Met Phe Phe Glu Ser Tyr Gly His Ser
2375 2380 2385
Ser His Met Leu Thr Asp Thr Lys Thr Gly Leu His Phe Asn Ile
2390 2395 2400
Asn Glu Ala Ile Glu Gln Gly Thr Ile Asp Lys Ala Leu Val Lys
2405 2410 2415
Lys Tyr Gln Glu Gly Leu Ile Thr Leu Thr Glu Leu Ala Asp Ser
2420 2425 2430
Leu Leu Ser Arg Leu Val Pro Lys Lys Asp Leu His Ser Pro Val
2435 2440 2445
Ala Gly Tyr Trp Leu Thr Ala Ser Gly Glu Arg Ile Ser Val Leu
2450 2455 2460
Lys Ala Ser Arg Arg Asn Leu Val Asp Arg Ile Thr Ala Leu Arg
2465 2470 2475
Cys Leu Glu Ala Gln Val Ser Thr Gly Gly Ile Ile Asp Pro Leu
2480 2485 2490
Thr Gly Lys Lys Tyr Arg Val Ala Glu Ala Leu His Arg Gly Leu
2495 2500 2505
Val Asp Glu Gly Phe Ala Gln Gln Leu Arg Gln Cys Glu Leu Val
2510 2515 2520
Ile Thr Gly Ile Gly His Pro Ile Thr Asn Lys Met Met Ser Val
2525 2530 2535
Val Glu Ala Val Asn Ala Asn Ile Ile Asn Lys Glu Met Gly Ile
2540 2545 2550
Arg Cys Leu Glu Phe Gln Tyr Leu Thr Gly Gly Leu Ile Glu Pro
2555 2560 2565
Gln Val His Ser Arg Leu Ser Ile Glu Glu Ala Leu Gln Val Gly
2570 2575 2580
Ile Ile Asp Val Leu Ile Ala Thr Lys Leu Lys Asp Gln Lys Ser
2585 2590 2595
Tyr Val Arg Asn Ile Ile Cys Pro Gln Thr Lys Arg Lys Leu Thr
2600 2605 2610
Tyr Lys Glu Ala Leu Glu Lys Ala Asp Phe Asp Phe His Thr Gly
2615 2620 2625
Leu Lys Leu Leu Glu Val Ser Glu Pro Leu Met Thr Gly Ile Ser
2630 2635 2640
Ser Leu Tyr Tyr Ser Ser
2645
<210> 2
<211> 452
<212> PRT
<213> 人工序列
<400> 2
Lys Thr Val Asn Leu Val Leu Lys Asn Thr Gln Ala Ala Glu Ala Leu
1 5 10 15
Val Lys Leu Tyr Glu Thr Lys Leu Cys Glu Glu Glu Ala Val Ile Ala
20 25 30
Asp Lys Asn Asn Ile Glu Asn Leu Ile Ser Thr Leu Lys Gln Trp Arg
35 40 45
Ser Glu Val Asp Glu Lys Arg Gln Val Phe His Ala Leu Glu Asp Glu
50 55 60
Leu Gln Lys Ala Lys Ala Ile Ser Asp Glu Met Phe Lys Thr Tyr Lys
65 70 75 80
Glu Arg Asp Leu Asp Phe Asp Trp His Lys Glu Lys Ala Asp Gln Leu
85 90 95
Val Glu Arg Trp Gln Asn Val His Val Gln Ile Asp Asn Arg Leu Arg
100 105 110
Asp Leu Glu Gly Ile Gly Lys Ser Leu Lys Tyr Tyr Arg Asp Thr Tyr
115 120 125
His Pro Leu Asp Asp Trp Ile Gln Gln Val Glu Thr Thr Gln Arg Lys
130 135 140
Ile Gln Glu Asn Gln Pro Glu Asn Ser Lys Thr Leu Ala Thr Gln Leu
145 150 155 160
Asn Gln Gln Lys Met Leu Val Ser Glu Ile Glu Met Lys Gln Ser Lys
165 170 175
Met Asp Glu Cys Gln Lys Tyr Ala Glu Gln Tyr Ser Ala Thr Val Lys
180 185 190
Asp Tyr Glu Leu Gln Thr Met Thr Tyr Arg Ala Met Val Asp Ser Gln
195 200 205
Gln Lys Ser Pro Val Lys Arg Arg Arg Met Gln Ser Ser Ala Asp Leu
210 215 220
Ile Ile Gln Glu Phe Met Asp Leu Arg Thr Arg Tyr Thr Ala Leu Val
225 230 235 240
Thr Leu Met Thr Gln Tyr Ile Lys Phe Ala Gly Asp Ser Leu Lys Arg
245 250 255
Leu Glu Glu Glu Glu Ile Lys Arg Cys Lys Glu Thr Ser Glu His Gly
260 265 270
Ala Tyr Ser Asp Leu Leu Gln Arg Gln Lys Ala Thr Val Leu Glu Asn
275 280 285
Ser Lys Leu Thr Gly Lys Ile Ser Glu Leu Glu Arg Met Val Ala Glu
290 295 300
Leu Lys Lys Gln Lys Ser Arg Val Glu Glu Glu Leu Pro Lys Val Arg
305 310 315 320
Glu Ala Ala Glu Asn Glu Leu Arg Lys Gln Gln Arg Asn Val Glu Asp
325 330 335
Ile Ser Leu Gln Lys Ile Arg Ala Glu Ser Glu Ala Lys Gln Tyr Arg
340 345 350
Arg Glu Leu Glu Thr Ile Val Arg Glu Lys Glu Ala Ala Glu Arg Glu
355 360 365
Leu Glu Arg Val Arg Gln Leu Thr Ile Glu Ala Glu Ala Lys Arg Ala
370 375 380
Ala Val Glu Glu Asn Leu Leu Asn Phe Arg Asn Gln Leu Glu Glu Asn
385 390 395 400
Thr Phe Thr Arg Arg Thr Leu Glu Asp His Leu Lys Arg Lys Asp Leu
405 410 415
Ser Leu Asn Asp Leu Glu Gln Gln Lys Asn Lys Leu Met Glu Glu Leu
420 425 430
Arg Arg Lys Arg Asp Asn Glu Glu Glu Leu Leu Lys Leu Ile Lys Gln
435 440 445
Met Glu Lys Asp
450

Claims (1)

1.一种非治疗、非诊断目的的检测大疱性类天疱疮BP230 IgE型抗体的方法，其特征在于，包括步骤：
a)将样品与多肽接触，所述多肽的氨基酸序列如SEQ ID NO.2所示，所述的多肽是和BP230 IgE型抗体结合；
b)检测是否形成抗原-抗体复合物，其中形成复合物就表示样品中存在大疱性类天疱疮BP230 IgE型抗体。
CN201910755415.2A 2019-08-15 2019-08-15 大疱性类天疱疮BP230 IgE抗体的抗原表位多肽及其用途 Active CN110437326B (zh)

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US20030138803A1 (en) * 2001-07-27 2003-07-24 Brooksbank Robert Alan Identification and use of molecules implicated in pain
CN101631877A (zh) * 2007-01-09 2010-01-20 百时美施贵宝公司 用于预测可与前列腺细胞中蛋白酪氨酸激酶和/或蛋白酪氨酸激酶途径相互作用和/或对其进行调节的化合物的活性的多核苷酸的鉴定
CN101333255B (zh) * 2008-07-04 2011-08-10 中国人民解放军第四军医大学 人源性抗大疱性类天疱疮抗原bp180单链抗体及其应用
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