title: Preparation of polyclonal antibodies for nateglinide (NTG) and development of a sensitive chemiluminescent immunoassay to detect NTG in tablets and serum.
abstract: In this study, we prepared polyclonal antibodies against anti-diabetic drug nateglinide (NTG), and established a sensitive chemiluminescent immunoassay (CLIA) to detect NTG in tablets and serum. Two kinds of immunogens were synthesized using ethylcarbodiimide (EDC)/hydroxysuccinimide (NHS) and carbonyldiimidazole (CDI)/4-dimethylaminopyridine (DMAP) as coupling reagents respectively. When activated by EDC/NHS, more molecules of NTG coupled with carrier protein in immunogens. A horseradish peroxidase (HRP)-luminol-H2O2 system with p-iodophenol enhancement was applied in the CLIA analysis. The antibodies in EDC/NHS group showed higher titer, sensitivity and wider detection linear range than those in CDI/DMAP group, and were chosen for next studies. The developed CLIA assay exhibited good selectivity towards NTG among structually similar analogs. The method could detect as low as 0.35 ng mL(-1) NTG in buffer, 2.1 ng mL(-1) NTG in serum and 0.84 ng mL(-1) NTG in tablets. The CLIA method provided consistent results with HPLC method (r=0.9986) in determination of NTG from 5.0 to 400 µg mL(-1). The CLIA method could detect 78 samples in one assay, and the samples need only dilution in pretreatment. As a summary, this research offers a sensitive assay for high-throughout screening of NTG in formulation control and pharmacokinetic studies.
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Zheng
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.affiliation: Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Tianjin, 300060, China; Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China.
authors.forename: Zhaorui
authors.initials: Z
authors.lastname: Song
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China.
authors.forename: Yaqing
authors.initials: Y
authors.lastname: Dong
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China.
authors.forename: Yufen
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Tianjin, 300060, China; Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China.
authors.forename: Zhongsheng
authors.initials: Z
authors.lastname: Tong
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China; Tianjin Sungene Biotech Co., Ltd., Tianjin 300450, China.
authors.forename: Chuan
authors.initials: C
authors.lastname: Deng
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China.
authors.forename: Yongmei
authors.initials: Y
authors.lastname: Yin
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China. Electronic address: mengmeng@nankai.edu.cn.
authors.forename: Meng
authors.initials: M
authors.lastname: Meng
authors.affiliation: College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, China.
authors.forename: Rimo
authors.initials: R
authors.lastname: Xi
mesh_terms: D000906:Antibodies / Q000276:immunology*; D001774:Blood Chemical Analysis; D002626:Chemistry, Pharmaceutical; D003510:Cyclohexanes / Q000032:analysis* / Q000097:blood / Q000276:immunology; D007118:Immunoassay / Q000379:methods*; D007700:Kinetics; D057230:Limit of Detection; D008163:Luminescent Measurements / Q000379:methods*; D000077715:Nateglinide; D010649:Phenylalanine / Q000031:analogs & derivatives* / Q000032:analysis / Q000097:blood / Q000276:immunology; D013607:Tablets
chemical_list: D000906:Antibodies; D003510:Cyclohexanes; D013607:Tablets; D000077715:Nateglinide; D010649:Phenylalanine
keywords: Chemiluminescence immunoassay; Nateglinide; Polyclonal antibodies; Serum; Tablets