Intrinsic structure of biological layers: Vertical inhomogeneity profiles characterized by label-free optical waveguide biosensors

When various analytes and functional layers are deposited on the surface of optical waveguide based biochips one faces difficulties in correctly interpreting the experimental data. For example, the deposited layer can be highly ordered (lipid bilayers, oriented receptors etc.) or can have significant inhomogeneity both perpendicular and parallel to the sensor surface (adsorbed polymer films, living cells). The generally applied simple optical model, which treats the deposited analyte layers homogeneous, isotropic and usually relatively thin compared to the wavelength of light, therefore, fails in most of the practical cases. In the present contribution, we systematically investigate the limitations of the widely applied optical models, when the analyte layer on the sensing surface has a vertically inhomogeneous refractive index profile. As examples of more realistic density profiles, the step-index, linear, exponential, power law and Gaussian refractive index distributions on various types of waveguide biochips are investigated using analytical and numerical model calculations. The limitations and the possible errors of the homogeneous thin adlayer model are pointed out. It is shown that for all of the vertically inhomogeneous profiles the refractive index obtained from the homogeneous thin layer modeling underestimates the true averaged refractive index of the layer. The calculated thickness can be over or underestimated, even taking up negative values in some cases. This behavior is similar to what was observed for positively birefringent thin adlayers treated with the homogeneous and isotropic model. It is also shown that the surface mass coverages calculated using the thickness and refractive index obtained from the homogeneous and isotropic modeling underestimate the real coverage values. The above errors are smallest when the reverse waveguide sensor design is applied to investigate vertically inhomogeneous analytes.