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

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

1,2,†
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.
Keywords: EDC; NHS; sulfoNHS; antibody crosslinking; APTES-functionalized platforms; ELISA; SPR EDC; NHS; sulfoNHS; antibody crosslinking; APTES-functionalized platforms; ELISA; SPR
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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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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