Karen M. Grace

Waveguide-Based Biosensors for Pathogen Detection

Optical phenomena such as fluorescence, phosphorescence, polarization, interference and non-linearity have been extensively used for biosensing applications. Optical waveguides (both planar and fiber-optic) are comprised of a material with high permittivity/high refractive index surrounded on all sides by materials with lower refractive indices, such as a substrate and the media to be sensed. This arrangement allows coupled light to propagate through the high refractive index waveguide by total internal reflection and generates an electromagnetic wave—the evanescent field—whose amplitude decreases exponentially as the distance from the surface increases. Excitation of fluorophores within the evanescent wave allows for sensitive detection while minimizing background fluorescence from complex, “dirty” biological samples. In this review, we will describe the basic principles, advantages and disadvantages of planar optical waveguide-based biodetection technologies. This discussion will include already commercialized technologies (e.g., Corning’s EPIC® Ô, SRU Biosystems’ BIND™, Zeptosense®, etc.) and new technologies that are under research and development. We will also review differing assay approaches for the detection of various biomolecules, as well as the thin-film coatings that are often required for waveguide functionalization and effective detection. Finally, we will discuss reverse-symmetry waveguides, resonant waveguide grating sensors and metal-clad leaky waveguides as alternative signal transducers in optical biosensing.

Pathogen detection using single mode planar optical waveguides

We utilize an optical waveguide-based biosensor recently developed at Los Alamos National Laboratory for the sensitive and specific detection of protein markers. Our planar optical waveguides are based on single mode structures that provide high optical field intensity at the active surface while providing discrimination from sample background fluorescence through spatial filtering. We have incorporated sandwich immunoassays where the capture antibody is immobilized on a biocompatible film attached to the surface of an optical waveguide. The approach can be adapted to any marker protein and is faster (<10 min), more sensitive, and as specific as conventional enzyme-linked immunosorbent assays (ELISA, the current lab standard). Results are presented for the detection of several analytes of Bacillus anthracis including protective antigen, a virulence marker for B. anthracis, and the bacterial cell or cellular debris.

Integrated optical biosensor for detection of multivalent proteins.

We have developed a simple, highly sensitive and specific optical waveguide sensor for the detection of multivalent proteins. The optical biosensor is based on optically tagged glycolipid receptors embedded within a fluid phospholipid bilayer membrane formed upon the surface of a planar optical waveguide. Binding of multivalent cholera toxin triggers a fluorescence resonance energy transfer that results in a two-color optical change that is monitored by measurement of emitted luminescence above the waveguide surface. The sensor approach is highly sensitive and specific and requires no additional reagents and washing steps. Demonstration of protein-receptor recognition by use of planar optical waveguides provides a path forward for the development of fieldable miniaturized biosensor arrays.

Thin-film chemical sensors with waveguide Zeeman interferometry

We describe a highly sensitive chemical sensor scheme using a channel waveguide with a selective surface coating based on polarimetric Zeeman interferometry. The sensing is based on measurement of the phase difference between TE and TM modes propagating in the anisotropic waveguide structure under exposure to toluene vapour. A real-time and reversible response at low ppm level is observed. Modelling results of the sensor structure to further increase its sensitivity are presented.

Reagentless optical biosensor

Critical to our ability to respond effectively to a biothreat attack is the development of sensitive and specific sensor systems that can easily be used for rapid screening of potential victims for infection due to biothreat agents and detection of pathogens in the environment. To help address these needs, we have developed a Reagentless Optical Biosensor (ROB) based on protein specific assays and waveguide-based evanescent fluorescence excitation. Modeled on host pathogen interactions, the sensors membrane based assay provides rapid, sensitive detection without the addition of reagents. We report here the development of two waveguide based detection systems: a laboratory sensor test-bed system and a handheld, battery operated, prototype. Evanescent fluorescence excitation using planar optical waveguides provides spatial filtering of background auto-fluorescence found in many natural samples, thereby permitting direct analysis of complex environmental and medical samples. The waveguide based assay is fully self-contained in a small, exchangeable cartridge that is optically coupled to the sensor detection system making ROB simple to use and offering the possibility of inexpensive, disposable sensor elements. Using assays for cholera toxin we compare results using flourimetry of vesicle solutions against results for our waveguide based test-bed and prototype sensor systems.

Integrated optical toxin sensor

We have developed a method for simple and highly sensitive detection of multivalent proteins using an optical waveguide sensor. The optical biosensor is based on optically tagged glycolipid receptors imbedded within a fluid phospholipid bilayer membrane formed on the surface of a planar optical waveguide. The binding of multivalent toxin initiates a fluorescence resonance energy transfer resulting in a distinctive spectral signature that is monitored by measuring emitted luminescence above the waveguide surface. The sensor methodology is highly sensitive and specific, and requires no additional reagents or washing steps. Demonstration of the utility of protein-receptor recognition using planar optical waveguides is shown here by the detection of cholera toxin.

Thin film waveguide sensors for chemical detection

A chemical sensor scheme, based on selective sensing surfaces and highly sensitive integrated optical transduction is presented. Self-assembly techniques are used to covalently attach species selective films onto the surface of silicon nitride waveguides. Exposure to targeted analytes results in selective absorption of these molecules onto the waveguide surface, causing a change in the effective refractive index of the guided modes. These relative changes in effective refractive indices of TM and TE modes are measured using Zeeman interferometry. The measurements demonstrate reversible, real time sensing of volatile organic compounds at ppm level. Improvements in the waveguide design are proposed to further increase the sensor performance.

Real time chemical detection using species selective thin films and waveguide Zeeman interferometry

We present a chemical sensor scheme based on selective sensing surfaces and highly sensitive integrated optical transduction methods. Using self-assembly techniques, species selective thin-films are covalently attached to the surface of Si3N4 channel waveguides to produce robust sensor elements. Exposure to targeted analytes results in the selective absorption of these molecules onto the waveguide surface causing a change in the effective refractive index of the guided modes. These relative changes in effective refractive index between TE and TM modes are precisely measured using Zeeman interferometry. Our measurements demonstrate reversible, real time sensing of volatile organic compounds at ppm levels.

In-situ chemical detection based on photonic devices

A novel scheme of a laser-based chemical sensor has been examined. The scheme is based on the lasing frequency shift of a DBR laser as a result of refractive index change of the sensitive coating in the presence of chemicals in question. The applicability and advantages of different schemes are discussed. The results of preliminary experiments related to the construction and stability of an external cavity DBR laser and interferometric measurements of refractive index change are presented.