No abstract

Optical methods based on recording a phase shift of electromagnetic waves offer high sensitivity for detection of bio-reactions. The method of Total Internal Reflection Ellipsometry (TIRE), which records simultaneously two parameters Ψ and ∆ related, respectively, to the amplitudes and phases of p-and s-components of polarized light, was exploited here for detection of mycotoxins (T-2, zearalanone, and aflatoxin). The TIRE direct label-free immunoassay enables detecting the above mycotoxins in sub-ppb concentrations in water as well as in food extracts. Further enhancement of sensitivity can be achieved with the use of SiO 2 /Si 3 N 4 /SiO 2 planar waveguides operating as polarization interferometers (PI).

No abstract

The method of spectroscopic ellipsometry in total internal reflection mode (TIRE) was utilised for detection of Aflatoxin B1. The method of TIRE with the improved data analysis was capable of detection of aflatoxin molecules in low concentrations (down to 0.04 ng/ml) using a label-free and cost-effective direct immunoassay format. TIRE study of the binding kinetics yielded a large value of the association constant in the range of 10 6 l mol −1 which is typical for highly specific immune reactions. The comparison of the experimental data for three mycotoxins studied (e.g. aflatoxin B1, T-2 mycotoxin, and zearalenone) confirmed a common mechanism of the sensitivity boost due to the aggregation of hydrophobic molecules of mycotoxins in aqueous solutions.

No abstract

A highly sensitive method of spectroscopic ellipsometry in total internal reflection mode (TIRE) was exploited for detecting β-amyloid peptide (Aβ(1-16)) in the direct immune reaction with monoclonal DE2 antibodies (raised against Aβ(1-16)) electrostatically immobilised on the surface of gold. A rapid detection of Aβ(1-16) in a wide range of concentrations from 5 μg/ml down to 0.05 ng/ml was achieved using a cost-effective and label-free direct immunoassay format. TIRE dynamic spectral measurements proved that the immune reaction between DE2 monoclonal antibodies and Aβ(1-16) is highly specific with the affinity constant K(D)=1.46×10(-8) mol/l. The same DE2 antibodies were utilised for detection of amyloid precursor protein APP(770), a larger protein containing Aβ(1-16) domain, using the quartz crystal microbalance (QCM) measurements in liquid. A combination of QCM and TIRE kinetics results allowed the evaluation of the originally unknown concentration of APP(770) in complete medium solution containing other proteins, salts, and amino acids.

This work describes a detailed quantitative interaction study between the novel plastidial chaperone receptor OEP61 and isoforms of the chaperone types Hsp70 and Hsp90 using the optical method of total internal reflection ellipsometry (TIRE). The receptor OEP61 was electrostatically immobilized on a gold surface via an intermediate layer of polycations. The TIRE measurements allowed the evaluation of thickness changes in the adsorbed molecular layers as a result of chaperone binding to receptor proteins. Hsp70 chaperone isoforms but not Hsp90 were shown to be capable of binding OEP61. Dynamic TIRE measurements were carried out to evaluate the affinity constants of the above reactions and resulted in clear discrimination between specific and nonspecific binding of chaperones as well as differences in binding properties between the highly similar Hsp70 isoforms.

No abstract