Investigating the operating mechanism of a diffraction based biosensor

Abstract

In this work, we describe our recent efforts aimed at determining the mechanism of signal change for a diffraction-based sensor (DBS) system. The DBS detects analyte-binding events by monitoring the change in diffraction efficiency that takes place when analyte molecules adsorb to target molecules that have been patterned onto a surface. The exact parameters that affect the intensity of the diffraction intensity are currently not well understood.In this work, the formalism used to describe the behaviour of volume-phase holography is used to understand the parameters that effect the diffraction intensity. It is hypothesized that the major factors that effect the diffraction intensity are the differences in optical path length between the wave trains that reflect off the diffraction grating and those that reffect off the substrate surface. Also key is the difference in refractive index between the two media. Two approaches were developed to investigate this hypothesis; the first was to develop a series of gratings of varying thickness using polyelectrolyte multilayers. The indices of refraction of these gratings were adjusted by the incorporation of charged gold nanoparticles. Since DBS systems operate by monitoring the binding of analyte molecules, a second series of experiments were developed to investigate the changes in diffraction intensity as micometer sized carboxylated beads were loaded onto an avidin grating. The first aspect that was investigated was the effect of adding more particles onto the grating surface on diffraction intensity. Second, the extent to which the particles reduced the periodicity of the diffraction grating, and the effect on the observed intensity of the diffraction signal were also investigated. Finally, this work shows the first use of a DBS system to extract the rate of and the maximum surface coverage of a specific binding reaction.