Antibody immobilization on to polystyrene substrate—on-chip immunoassay for horse IgG based on fluorescence |
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Authors: | Farzana Darain Kai Ling Gan Swee Chuan Tjin |
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Institution: | (1) Biomedical Engineering Research Centre, Nanyang Technological University, Singapore, 639553, Singapore;(2) Photonics Research Centre, School of Electrical and Electronic Engineering, Nanyang Technological University, Singapore, 639798, Singapore |
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Abstract: | A simple microfluidic immunoassay card was developed based on polystyrene (PS) substrate for the detection of horse IgG, an
inexpensive model analyte using fluorescence microscope. The primary antibody was captured onto the PS based on covalent bonding
via a self-assembled monolayer (SAM) of thiol to pattern the surface chemistry on a gold-coated PS. The immunosensor chip layers
were fabricated from sheets by CO2 laser ablation. The functionalized PS surfaces after each step were characterized by contact angle measurement, X-ray photoelectron
spectroscopy (XPS), and atomic force microscopy (AFM). After the antibody–antigen interaction as a sandwich immunoassay with
a fluorescein isothiocyanate (FITC)-conjugated secondary antibody, the intensity of fluorescence was measured on-chip to determine
the concentration of the target analyte. The present immunosensor chip showed a linear response range for horse IgG between
1 μg/ml and 80 μg/ml (r = 0.971, n = 3). The detection limit was found to be 0.71 μg/ml. The developed microfluidic system can be extended for various applications
including medical diagnostics, microarray detection and observing protein–protein interactions. |
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Keywords: | Polystyrene Covalent bonding Microfluidic card Fluorescence Horse IgG |
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