Generation and characterization of monoclonal antibodies that recognize human and murine supervillin protein isoforms.
Generation and characterization of monoclonal antibodies that recognize human and murine supervillin protein isoforms.
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识别人和鼠类超级蛋白质蛋白同工型的单克隆抗体的产生和表征。
DOI：
10.1371/journal.pone.0205910
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发表时间：
2018
期刊：
影响因子：
3.7
通讯作者：
Luna EJ
中科院分区：
文献类型：
--
作者：
Smith TC;Saul RG;Barton ER;Luna EJ
超级绒毛蛋白亚型与细胞增殖、肌动蛋白丝为基础的运动过程、囊泡运输和信号转导有关。然而，由于难以获得针对这些高度保守的膜相关蛋白的特异性抗体，对这些蛋白在癌症转移和生理过程中的作用的理解受到限制。为了便于研究超级绒毛蛋白的生物学功能，我们制备了针对细菌表达的人超级绒毛蛋白n端的单克隆抗体。两种兔Fc结构域嵌合单克隆抗体（克隆1E2/CPTC-SVIL-1； 4A8/CPTC-SVIL-2）和两种小鼠单克隆抗体（克隆5A8/CPTC-SVIL-3； 5G3/CPTC-SVIL-4）通过免疫印迹、免疫沉淀、免疫荧光显微镜和免疫组织化学染色对其结合位点、亲和力和有效性进行了表征。两种抗体（1E2, 5G3）识别仅在灵长类超级绒毛蛋白中发现的序列，而另外两种抗体（4A8, 5A8）特异性识别更广泛保守的构象表位。在这里确定的固定条件下，所有抗体在免疫印迹、免疫沉淀和免疫荧光显微镜下起作用。我们还发现5A8抗体在免疫组织学切片上起作用。这些抗体将为哺乳动物超级病毒的研究提供有用的工具。
Supervillin isoforms have been implicated in cell proliferation, actin filament-based motile processes, vesicle trafficking, and signal transduction. However, an understanding of the roles of these proteins in cancer metastasis and physiological processes has been limited by the difficulty of obtaining specific antibodies against these highly conserved membrane-associated proteins. To facilitate research into the biological functions of supervillin, monoclonal antibodies were generated against the bacterially expressed human supervillin N-terminus. Two chimeric monoclonal antibodies with rabbit Fc domains (clones 1E2/CPTC-SVIL-1; 4A8/CPTC-SVIL-2) and two mouse monoclonal antibodies (clones 5A8/CPTC-SVIL-3; 5G3/CPTC-SVIL-4) were characterized with respect to their binding sites, affinities, and for efficacy in immunoblotting, immunoprecipitation, immunofluorescence microscopy and immunohistochemical staining. Two antibodies (1E2, 5G3) recognize a sequence found only in primate supervillins, whereas the other two antibodies (4A8, 5A8) are specific for a more broadly conserved conformational epitope(s). All antibodies function in immunoblotting, immunoprecipitation and in immunofluorescence microscopy under the fixation conditions identified here. We also show that the 5A8 antibody works on immunohistological sections. These antibodies should provide useful tools for the study of mammalian supervillins.
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影响因子：
--
作者：
DeNardo, DG;Kim, HT;Brown, PH
通讯作者：
Brown, PH
影响因子：
3.7
作者：
Burridge K;Guilluy C
通讯作者：
Guilluy C
影响因子：
--
作者：
Di Domenico M;Giordano A
通讯作者：
Giordano A
DOI：
10.1073/pnas.0533805100
发表时间：
2003-03-04
影响因子：
11.1
作者：
Hedenfalk, I;Ringnér, M;Trent, J
通讯作者：
Trent, J
影响因子：
4.8
作者：
Fang, Zhiyou;Luna, Elizabeth J.
通讯作者：
Luna, Elizabeth J.