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Diagnostics 2012, 2(3), 23-33; https://doi.org/10.3390/diagnostics2030023

Comparison of 1-Ethyl-3-(3-Dimethylaminopropyl) Carbodiimide Based Strategies to Crosslink Antibodies on Amine-Functionalized Platforms for Immunodiagnostic Applications

1
Centre for Bioanalytical Sciences, National Centre for Sensor Research, Dublin City University, Dublin 9, Ireland
2
Bristol-Myers Squibb (BMS), Swords Laboratories, Watery Lane, Swords, Co. Dublin, Ireland
Current address: HSG-IMIT—Institut für Mikro-und Informationstechnik, Georges-Koehler-Allee 103, 79110 Freiburg, Germany
Received: 13 July 2012 / Revised: 14 August 2012 / Accepted: 22 August 2012 / Published: 27 August 2012
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Abstract

1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) alone, and in combination with N-hydroxysuccinimide (NHS) or sulfoNHS were employed for crosslinking anti-human fetuin A (HFA) antibodies on 3-aminopropyltriethoxysilane (APTES)-functionalized surface plasmon resonance (SPR) gold chip and 96-well microtiter plate. The SPR immunoassay and sandwich enzyme linked immunosorbent immunoassay (ELISA) for HFA clearly demonstrated that EDC crosslinks anti-HFA antibodies to APTES-functionalized bioanalytical platforms more efficiently than EDC/NHS and EDC/sulfoNHS at a normal pH of 7.4. Similar results were obtained by sandwich ELISAs for human Lipocalin-2 and human albumin, and direct ELISA for horseradish peroxidase. The more efficient crosslinking of antibodies by EDC to the APTES-functionalized platforms increased the cost-effectiveness and analytical performance of our immunoassays. This study will be of wide interest to researchers developing immunoassays on APTES-functionalized platforms that are being widely used in biomedical diagnostics, biosensors, lab-on-a-chip and point-of-care-devices. It stresses a critical need of an intensive investigation into the mechanisms of EDC-based amine-carboxyl coupling under various experimental conditions. View Full-Text
Keywords: EDC; NHS; sulfoNHS; antibody crosslinking; APTES-functionalized platforms; ELISA; SPR EDC; NHS; sulfoNHS; antibody crosslinking; APTES-functionalized platforms; ELISA; SPR
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Vashist, S.K. Comparison of 1-Ethyl-3-(3-Dimethylaminopropyl) Carbodiimide Based Strategies to Crosslink Antibodies on Amine-Functionalized Platforms for Immunodiagnostic Applications. Diagnostics 2012, 2, 23-33.

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