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Quantitative enzyme-linked immunosorbent assay (ELISA) for non-enzymatically glycated serum protein
Authors:H Nakayama  Z Makita  M Kato  S Taneda  H Yoshida  K Yanagisawa  S Nakagawa
Affiliation:1. Department of Electrical and Electronic Engineering, Chittagong University of Engineering and Technology, Chittagong, 4349, Bangladesh;2. Institute for Photonics and Advanced Sensing (IPAS) and School of Physical Sciences, The University of Adelaide, Adelaide, SA, 5005, Australia;3. ARC Centre of Excellence for Nanoscale Biophotonics, The University of Adelaide, Adelaide, SA, 5005, Australia;4. Department of Electrical and Computer Engineering, Purdue University, West Lafayette, IN, 47906, USA;5. Department of Electrical and Computer Engineering, University of Waterloo, Canada;1. Department of General, Visceral and Vascular Surgery, Charité – Universitätsmedizin Berlin, Berlin, Germany;2. Center for Diagnostic and Interventional Radiology and Nuclear Medicine, Charité – Universitätsmedizin Berlin, Berlin, Germany;3. Berlin Brandenburg School of Regenerative Therapies, Charité – Universitätsmedizin Berlin, Berlin, Germany;4. Laboratory of Multivariate Analysis and Global Modelling, Samara State Technical University, Samara, Russia;5. art photonics GmbH, Berlin, Germany;1. School of Chemistry and Chemical Engineering, Guangxi University for Nationalities, Key Laboratory of Guangxi Colleges and Universities for Food Safety and Pharmaceutical Analytical Chemistry, Guangxi Key Laboratory of Chemistry and Engineering of Forest Products, Guangxi Colleges and Universities Key Laboratory of Utilization of Microbial and Botanical Resources, Nanning, Guangxi 530008, China;2. School of Chemistry and Chemical Engineering, Guangxi University, Nanning, Guangxi 530004, China;1. Institute of Natural Medicinal Chemistry & Green Chemistry, Department of Pharmaceutical Engineering, School of Light Industry and Chemical Engineering, Guangdong University of Technology, Guangzhou 510006, People’s Republic of China;2. School of Chemistry & Environmental Engineering, Wuyi University, Jiangmen 529020, People’s Republic of China;3. Infinitus (China) Company Ltd, People’s Republic of China
Abstract:A competitive ELISA for quantitative determination of glucitollysine, the reduced hexose alcohol form of glucose conjugated to the epsilon amino group of lysine was developed. We applied it to measure non-enzymatically glycated serum proteins. The antiserum obtained by immunizing guinea pigs with reductively glycated human albumin was capable of identifying and quantitating glucitollysine residues of serum proteins in normal and diabetic subjects after reduction of the proteins with sodium borohydride. The ELISA assay developed here had satisfactory reproducibility as judged by the intra-assay precision of 2.3-7.6% and the interassay precision of 6.7-9.8%. Results from this assay procedure correlated well with those from the radioimmunoassay and the boronate affinity chromatography procedure. The data suggested that diabetic serum proteins contained at least three times as much immunochemically detectable glucitollysine residues as normal serum proteins after reduction of the proteins with sodium borohydride. This method allows to quantitate glucitollysine residues on any of the proteins that have been implicated in the pathological sequelae of diabetes.
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