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Dual-polarization interferometry (DPI) is an analytical technique that probes molecular layers adsorbed to the surface of a waveguide using the
evanescent wave In electromagnetics, an evanescent field, or evanescent wave, is an oscillating electric and/or magnetic field that does not propagate as an electromagnetic wave but whose energy is spatially concentrated in the vicinity of the source (oscillat ...
of a
laser A laser is a device that emits light through a process of optical amplification based on the stimulated emission of electromagnetic radiation. The word "laser" is an acronym for "light amplification by stimulated emission of radiation". The ...
beam. It is used to measure the
conformational change In biochemistry, a conformational change is a change in the shape of a macromolecule, often induced by environmental factors. A macromolecule is usually flexible and dynamic. Its shape can change in response to changes in its environment or oth ...
in proteins, or other biomolecules, as they function (referred to as the
conformation activity relationship Conformation generally means structural arrangement and may refer to: * Conformational isomerism, a form of stereoisomerism in chemistry ** Carbohydrate conformation ** Cyclohexane conformation ** Protein conformation ** Conformation activity rela ...
).


Instrumentation

DPI focuses laser light into two waveguides. One of these functions as the "sensing" waveguide having an exposed surface while the second one functions to maintain a reference beam. A two-dimensional interference pattern is formed in the far field by combining the light passing through the two waveguides. The DPI technique rotates the polarization of the laser, to alternately excite two polarization modes of the waveguides. Measurement of the interferogram for both polarizations allows both the
refractive index In optics, the refractive index (or refraction index) of an optical medium is a dimensionless number that gives the indication of the light bending ability of that medium. The refractive index determines how much the path of light is bent, ...
and the thickness of the adsorbed layer to be calculated. The polarization can be switched rapidly, allowing real-time measurements of
chemical reaction A chemical reaction is a process that leads to the chemical transformation of one set of chemical substances to another. Classically, chemical reactions encompass changes that only involve the positions of electrons in the forming and breaking ...
s taking place on a chip surface in a flow-through system. These measurements can be used to infer conformational information about the molecular interactions taking place, as the molecule size (from the layer thickness) and the fold density (from the RI) change. DPI is typically used to characterize
biochemical Biochemistry or biological chemistry is the study of chemical processes within and relating to living organisms. A sub-discipline of both chemistry and biology, biochemistry may be divided into three fields: structural biology, enzymology an ...
interactions by quantifying any
conformational change In biochemistry, a conformational change is a change in the shape of a macromolecule, often induced by environmental factors. A macromolecule is usually flexible and dynamic. Its shape can change in response to changes in its environment or oth ...
at the same time as measuring reaction rates, affinities and thermodynamics. The technique is quantitative and real-time (10 Hz) with a dimensional resolution of 0.01 nm. Extensions of dual-polarization interferometry also exist, namely multiple pathlength dual-polarization interferometry (MPL-DPI) and absorption enhanced DPI. In MPL-DPI quantification of both layer thickness and refractive index (density) and therefore mass per unit area can be made for in situ and ex-situ coated films, where normal DPI can only calculate film properties if the interferogram is constantly monitored. Absorption enhanced DPI (AE-DPI) is used to separate the mass of different molecules on the surface, exploiting the absorption of one of the molecular species compared to the other species on the surface.


Applications

A novel application for dual-polarization interferometry emerged in 2008, where the intensity of light passing through the waveguide is extinguished in the presence of crystal growth. This has allowed the very earliest stages in protein crystal nucleation to be monitored. Later versions of dual-polarization interferometers also have the capability to quantify the order and disruption in birefringent thin films. This has been used, for example, to study the formation of lipid bilayers and their interaction with membrane proteins.


References


Further reading

* * * * {{Protein structural analysis Biosensors Interferometry Protein–protein interaction assays Protein methods Polyelectrolytes Scientific instruments Scientific equipment