Fluorescence IgG immunosensor based on a micro flow cell containing controlled pore glass as immobilisation support

Abstract

Biosensor miniaturisation often requires the construction of micro volume cells using micro-machining techniques. In this work, a micro flow cell made of a transparent polymer [poly(methyl methacrylate)] developed for using with a fluorescence immunoglobulin G (IgG) immunosensor is described. The micro flow cell provides space to enclose a small amount of controlled pore glass (CPG), a transparent support that can host and covalently bind the biomolecules. The immobilisation of IgG on CPG permits the development of an optical immunosensor for the detection of fluorescein isothiocyanate (FITC)-labelled anti-IgG. In this immunosensor the excitation light is provided by an argon ion laser and guided by an optical fibre to the flow cell, where the fluorescence signal is filtered by a long-pass barrier filter (OG515) and then detected by a close positioned photodiode. This signal was found to be proportional to the amount of anti-IgG–FITC bound to the immobilised IgG during a direct immunochemical reaction. Characterisation of the CPG as an optical medium and immobilisation support was performed. CPG produces intense light scattering and good permeability to fluids, and also a typical immobilisation rate for IgG of about 90% of the initial amount of antibody. The described immunosensor shows a detection limit for anti-IgG–FITC of 6.3 nM and a sensitivity of 9.5 μV nM⁻¹. This immunoptode developed with a micro flow cell has been shown to be a suitable system for the detection of immunoglobulins.