title: Detection of antibodies against customized epitope: use of a coating antigen employing VEGF as fusion partner.
abstract: Diagnosis of many infectious, autoimmune diseases and cancers depends on the detection of specific antibodies against peptide epitope by enzyme-linked immunosorbent assay (ELISA). However, small peptides are difficult to be coated on the plate surfaces. In this study, we selected GnRH as a model hapten to evaluate whether VEGF121 would be suitable as an irrelevant hapten-carrier to develop a universal platform for specific antibodies detection. Firstly, GnRH was fused to the C terminus of VEGF121 and the resultant fusion protein VEGF-GnRH expressed effectively as inclusion bodies in Escherichia coli. Thereafter, VEGF-GnRH was easily purified to near homogeneity with a yield of about 235 mg from 2.1 L induced culture. At last, VEGF-GnRH was used to perform ELISA and western blot, and our results suggested that VEGF-GnRH was capable of detecting anti-GnRH antibodies in sera both qualitatively and quantitatively. Indeed, previous studies of our laboratory had demonstrated that other fusion proteins such as VEGF-Aβ10, VEGF-GRP, VEGF-CETPC, and VEGF-βhCGCTP37 were able to detect their corresponding antibodies specifically. Therefore, VEGF121 may be a suitable irrelevant fusion partner of important diagnostic peptide markers. Our works would shed some light on the development of a universal platform for detection of specific antibodies.
authors.affiliation: Key Laboratory of Human Functional Genomics of Jiangsu Province, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Nanjing Medical University, Nanjing, 210029, China, wangxuejun@njmu.edu.cn.
authors.forename: Xue Jun
authors.initials: XJ
authors.lastname: Wang
authors.forename: Li Jun
authors.initials: LJ
authors.lastname: Zhou
authors.forename: Xue Jiang
authors.initials: XJ
authors.lastname: Zhu
authors.forename: Kai
authors.initials: K
authors.lastname: Gu
authors.forename: Jie
authors.initials: J
authors.lastname: Wu
authors.forename: Tai Ming
authors.initials: TM
authors.lastname: Li
authors.forename: Li
authors.initials: L
authors.lastname: Yuan
authors.forename: Rong Yue
authors.initials: RY
authors.lastname: Cao
mesh_terms: D000818:Animals; D000906:Antibodies / Q000032:analysis* / Q000276:immunology; D000941:Antigens / Q000235:genetics / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000295:instrumentation* / Q000379:methods; D000939:Epitopes / Q000235:genetics / Q000276:immunology*; D007987:Gonadotropin-Releasing Hormone / Q000235:genetics / Q000276:immunology*; D007114:Immunization; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000276:immunology*
chemical_list: D000906:Antibodies; D000941:Antigens; D000939:Epitopes; D011993:Recombinant Fusion Proteins; D042461:Vascular Endothelial Growth Factor A; D007987:Gonadotropin-Releasing Hormone