Generation of monoclonal antibodies and development of an immunofluorometric assay for the detection of CUZD1 in tissues and biological fluids
Generation of monoclonal antibodies and development of an immunofluorometric assay for the detection of CUZD1 in tissues and biological fluids
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DOI：
10.1016/j.clinbiochem.2017.07.012
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发表时间：
2017-12-01
影响因子：
2.8
通讯作者：
Blasutig, Ivan M.
中科院分区：
文献类型：
--
作者：
Farkona, Sofia;Soosaipillai, Antoninus;Blasutig, Ivan M.
背景资料：CUB和含透明质酸样结构域蛋白1（CUZD 1）被鉴定为胰腺特异性蛋白，并被提议作为胰腺相关疾病的候选生物标志物。组织和生物体液中的CUZD 1蛋白水平尚未得到广泛研究。本研究的目的是产生针对CUZD 1的特异性抗体，以评估CUZD 1在组织和生物fluids.Methods表达：产生小鼠单克隆抗体对CUZD 1进行免疫组化分析，并开发一种敏感和特异性的酶联免疫吸附试验（ELISA）。CUZD 1蛋白的表达进行了评估，在各种人体组织提取物和生物液体和凝胶过滤色谱衍生馏分的胰腺组织提取物，胰液和重组protein.Results：免疫组化染色的CUZD 1在胰腺组织中显示，该蛋白是本地化的腺泡细胞和腔的腺泡。Western blot检测胰腺组织提取液和胰液中的蛋白质。新开发的ELISA在胰腺中测量了高水平的CUZD 1，在其他几个组织中测量了低得多但可检测的水平。在测试的生物体液中，CUZD 1表达仅在胰液中检测到。胰腺组织提取物，胰液和重组CUZD 1的凝胶过滤色谱衍生馏分的分析表明，该蛋白存在于高分子量的蛋白complex.Conclusion：本研究描述了针对CUZD 1蛋白，其组织表达模式和水平在几种生物液体的工具的发展。这些新工具将有助于未来的研究，旨在描绘CUZD 1在生理学和病理学中的作用。
Background: CUB and zona pellucida-like domain-containing protein 1 (CUZD1) was identified as a pancreas-specific protein and was proposed as a candidate biomarker for pancreatic related disorders. CUZD1 protein levels in tissues and biological fluids have not been extensively examined. The purpose of the present study was to generate specific antibodies targeting CUZD1 to assess CUZD1 expression within tissues and biological fluids.Methods: Mouse monoclonal antibodies against CUZD1 were generated and used to perform immunohistochemical analyses and to develop a sensitive and specific enzyme-linked immunosorbent assay ( ELISA). CUZD1 protein expression was assessed in various human tissue extracts and biological fluids and in gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant protein.Results: Immunohistochemical staining of CUZD1 in pancreatic tissue showed that the protein is localized to the acinar cells and the lumen of the acini. Western blot analysis detected the protein in pancreatic tissue extract and pancreatic juice. The newly developed ELISA measured CUZD1 in high levels in pancreas and in much lower but detectable levels in several other tissues. In the biological fluids tested, CUZD1 expression was detected exclusively in pancreatic juice. The analysis of gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant CUZD1 suggested that the protein exists in high molecular weight protein complexes.Conclusion: This study describes the development of tools targeting CUZD1 protein, its tissue expression pattern and levels in several biological fluids. These new tools will facilitate future investigations aiming to delineate the role of CUZD1 in physiology and pathobiology.