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Efficient Detection of Pre-Proinsulin by Double Antibody Sandwich ELISA

文献类型：期刊论文

作者Zhu, Zhu1,2,3; Wang, Han1; Wang, Li1; Wei, Zhou2; Zheng, Zhiming1; Wang, Peng1,3
刊名APPLIED SCIENCES-BASEL
出版日期2022-10-01
卷号12
关键词recombinant human insulin pre-proinsulin monoclonal antibody sandwich ELISA
DOI10.3390/app12199868
通讯作者Zheng, Zhiming(zhengzhiming2014@163.com) ; Wang, Peng(pengwang@ipp.ac.cn)
英文摘要To detect residual pre-proinsulin (PPI) in recombinant human insulin production, an analytical method based on double-antibody sandwich ELISA was developed in this study. The BALB/c mice were immunized with PPI, and the hybridomas secreting anti-PPI monoclonal antibodies were obtained using the conventional cell fusion technique and ELISA screening. We purified the antibody using a Protein G gel column and identified its purity by SDS-PAGE. The sandwich ELISA was used to explore the pairing effect, and the specificity of the paired antibody was determined. We selected a paired antibody with relatively good specificity to establish sandwich ELISA, constructed a quantitative curve, and evaluated the accuracy and sensitivity of the method. Six anti-PPI monoclonal antibodies were obtained, named P1, P2, P3, P4, P5 and P6, of which P5 had the highest titer value. The sandwich ELISA method was established with P5 for plating and P2 as detection antibodies. The linear range of the quantitative curve of PPI by sandwich ELISA was 0.645 to 82.5 pg/mL, the recovery was 95% and the detection limit was 3.06 pg/mL. In this study, we prepared six anti-PPI monoclonal antibodies and established the sandwich ELISA method to detect PPI in process control and product release control for recombinant human insulin production.
WOS关键词PURIFICATION PROCESS ; DIABETES-MELLITUS ; INSULIN ; OBESITY
资助项目Major Projects of Science and Technology of Anhui Province[202103a06020003] ; China National Key Research and Development Program[2019YFA0904300] ; China National Key Research and Development Program[2019YFA0904304]
WOS研究方向Chemistry ; Engineering ; Materials Science ; Physics
语种英语
出版者MDPI
WOS记录号WOS:000866632300001
资助机构Major Projects of Science and Technology of Anhui Province ; China National Key Research and Development Program
源URL[http://ir.hfcas.ac.cn:8080/handle/334002/129325]  
专题中国科学院合肥物质科学研究院
通讯作者Zheng, Zhiming; Wang, Peng
作者单位1.Chinese Acad Sci, Hefei Inst Phys Sci, Hefei 230031, Peoples R China
2.Hefei Tianmai Biotechnol Dev Co Ltd, Hefei 230061, Peoples R China
3.Univ Sci & Technol China, Sci Isl Branch, Grad Sch, Hefei 230026, Peoples R China
推荐引用方式
GB/T 7714		 Zhu, Zhu,Wang, Han,Wang, Li,et al. Efficient Detection of Pre-Proinsulin by Double Antibody Sandwich ELISA[J]. APPLIED SCIENCES-BASEL,2022,12.
APA Zhu, Zhu,Wang, Han,Wang, Li,Wei, Zhou,Zheng, Zhiming,&Wang, Peng.(2022).Efficient Detection of Pre-Proinsulin by Double Antibody Sandwich ELISA.APPLIED SCIENCES-BASEL,12.
MLA Zhu, Zhu,et al."Efficient Detection of Pre-Proinsulin by Double Antibody Sandwich ELISA".APPLIED SCIENCES-BASEL 12(2022).

入库方式： OAI收割

来源：合肥物质科学研究院

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