Antibody immobilization on to polystyrene substrate—on-chip immunoassay for horse IgG based on fluorescence

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 CO(2) 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 microg/ml and 80 microg/ml (r = 0.971, n = 3). The detection limit was found to be 0.71 microg/ml. The developed microfluidic system can be extended for various applications including medical diagnostics, microarray detection and observing protein-protein interactions.

A bead-based fluorescence immunosensing technique enabled by the integration of Förster resonance energy transfer and optoelectrokinetic concentration

“…A LDL concentration of 5 nM was eventually achieved. Compared with conventional substrate-based immunosensors, [36][37][38][39] this approach is more flexible and can be extended to different measurements when particles are conjugated with different antibodies. As a result, the bead-based fluorescence immunosensing enabled by the optoelectrokinetic technique provides insight to the possibility of developing early diagnoses for more other diseases.…”

Solvent effect on polystyrene surface roughness on top of QCM sensor

“…Polystyrene material was chosen not only because of its glassy property but also its property which can be used to immobilized biomolecule [4,[5][6][7][8]. Polystyrene has been used for a long time as ELISA plate where biomolecule under investigation was immobilized on the polystyrene surface plate.…”

Immobilization and trapping of living bacteria and applications in corrosion studies

“…Depending on the characteristics of the surface, these molecules contain reactive groups, such as − SH, − COOH, − NH 2 , or − OH, which in turn are used to graft antibodies to the surface either directly or indirectly (i.e., through linker, or tether, molecules). In particular, the use of heterobifunctional molecules as tether molecules with functional groups on both sides has become important in recent years (Bhatia et al ., 1989, Darain et al ., 2009Kausaite-Minkstimiene et al ., 2010 ;Rezania et al ., 1999 ;Shriver-Lake et al ., 1997 ). Unlike direct attachment, a tether molecule targets the amino acid residues located within the lower portion of an antibody for covalent attachment (Shriver-Lake et al ., 1997 ).…”