抗猪囊尾蚴IgG4抗体的特异性抗原筛选

中国寄生虫学与寄生虫病杂志 ›› 2017, Vol. 35 ›› Issue (5): 509-511.

抗猪囊尾蚴IgG4抗体的特异性抗原筛选

李瑾¹, 王燕², 魏庆宽¹, 贾凤菊¹, 肖婷¹, 孙慧¹, 于振华¹, 黄炳成^1,^*()

1 山东省寄生虫病防治研究所,世界卫生组织淋巴丝虫病和绦/囊虫病合作中心,济宁 272033
2 复旦大学附属妇产科医院研究所,上海 200011

收稿日期:2017-04-11 出版日期:2017-10-30 发布日期:2018-01-08
通讯作者: 黄炳成
基金资助:
山东省自然科学基金（No. ZR2015YL033,ZR2015YL051）;国家自然科学基金（No. 30872208）;山东省医学科学院医药卫生科技创新工程

Screening of antigens specific for the anti-Cysticercus cellulosae IgG4 antibody

Jin LI¹, Yan WANG², Qing-kuan WEI¹, Feng-ju JIA¹, Ting XIAO¹, Hui SUN¹, Zhen-hua YU¹, Bing-cheng HUANG^1,^*()

1 Shandong Institute of Parasitic Diseases, WHO collaborating Center for Lymphatic Filariasis and Taeniasis/Cysticercosis, Jining, 272033, China
2 Obstetrics and Gynecology Hospital of Fudan University, Shanghai 200011, China

Received:2017-04-11 Online:2017-10-30 Published:2018-01-08
Contact: Bing-cheng HUANG
Supported by:
Supported by the Natural Science Fund of Shandong Province(No. ZR2015YL033, ZR2015YL051), the National Natural Science Foundation of China(No. 30872208)and the Innovation Project of Shandong Academy of Medical Sciences

摘要/Abstract

摘要：

将囊液抗原蛋白采用双向凝胶电泳分离,经考马斯亮蓝染色后用双向电泳图像分析软件分析蛋白点,以脑囊尾蚴病患者血清为一抗进行蛋白质印迹分析（Western blotting）,选取阳性蛋白点进行质谱分析,获取肽质量指纹图谱,采用Mascot软件在NCBI中搜寻、比对,筛选出抗猪囊尾蚴IgG4特异性抗原。囊液抗原经双向电泳检测到（201 ± 5）个蛋白斑点,相对分子质量（M_r）为10 000~170 000,等电点（pI）为3.0~10.0。Western blotting分析显示,与患者血清中特异IgG4抗体结合的抗原以小分子量蛋白为主,从中筛选出51个（抗原）点进行质谱分析,其中有21个点峰值大于39。对比分析显示,高峰值点与10种囊尾蚴蛋白有关,其中最为密切相关的为4种蛋白,按分值高低排序分别为Ts8B2、Ts8B3、10ku抗原和Sequence 3。

关键词: 猪囊尾蚴, 囊液抗原, IgG4, 双向电泳

Abstract:

The prepared cyst fluid antigen was separated by two-dimensional electrophoresis(2-DE). The protein spots were analyzed by ImageMaster 2D Plantinum 6.0 software after Coomassie blue staining, on which Western blotting analysis was performed using sera of patients with cerebral cysticercosis as the primary antibody. The positive protein spots were selected and analyzed by Matrix-assisted laser desorption ionization tandem mass spectrometry (GC-MS), and the peptide mass fingerprints were obtained. The specific Cysticercus cellulosae antigen for IgG4 antibody was identified with the Mascot software by searching and alignment in NCBI. 2-DE detected 201 ± 5 protein spots in the cystic acid antigen, with a relative molecular weight （M_r） of 10 000-170 000 and a pI of 3.0-10.0. Western blotting analysis showed that the antigens binding the specific IgG4 antibody were mainly small molecular-weight proteins. Fifty-one(antigen) spots were obtained from them for mass spectrometry, and 21 of them were positive(peak score>39). The positive spots were related with 10 cysticercus proteins, most closely with Ts8B2, Ts8B3, 10ku antigen and Sequence 3.

Key words: Cysticercus cellulosae, Cysticercus fluid antigens, IgG4, Two-dimensional electrophoresis

中图分类号:

R532.32

李瑾, 王燕, 魏庆宽, 贾凤菊, 肖婷, 孙慧, 于振华, 黄炳成. 抗猪囊尾蚴IgG4抗体的特异性抗原筛选[J]. 中国寄生虫学与寄生虫病杂志, 2017, 35(5): 509-511.

Jin LI, Yan WANG, Qing-kuan WEI, Feng-ju JIA, Ting XIAO, Hui SUN, Zhen-hua YU, Bing-cheng HUANG. Screening of antigens specific for the anti-Cysticercus cellulosae IgG4 antibody[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2017, 35(5): 509-511.

图/表 2

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蛋白名称	NCBI检索ID	相对分子质量	等电点	分值
RS1	gi22657787	9217	8.9	45
Secreted antigen Ts8B3	gi23477226	7570	6.71	105
Cysticercosis 10ku antigen	gi112785160	9650.2	8.89	100
Secreted antigen Ts8B2	gi23477224	9680.2	9.15	115
Sequence 3 from patent US 6589752	gi33757615	9576.2	6.58	85
8 ku diagnostic antigen TsRS1	gi19879956	8641.7	8.91	41
Unnamed protein product	gi13185479	8271.4	8.9	41
Immunogenic protein Ts21	gi7339851	9671.3	9.23	40
Immunogenic protein	gi12802157	9458.2	9.23	40
RS1 variant	gi112819965	9589.2	9.22	39