Evanescent Wave

Xingwei Wang

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Optical fiber

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Reflection/Refraction

If θ < θc, as with the red ray in the above figure, the ray will split. Some of the ray will reflect off the boundary, and some will refract as it passes through. If θ > θc, as with the blue ray, all of the ray reflects from the boundary. None passes through.

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Snell's law

1 1 2 2s i n s i nn nθ θ=

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Critical angle

The minimum angle of incidence at which total internal reflection occurs n2 is the refractive index of the less dense medium, and n1 is the refractive index of the denser medium

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arcsincnn

θ⎛ ⎞

= ⎜ ⎟⎝ ⎠

Plot of angles (θreflection; θrefraction)The media are quartz (n1 = 1.46) and water (n2 = 1.33).

Evanescent Wave

θ > θc, some incident energy penetrate into the second medium. Forms an electromagnetic field that oscillates with the same frequency as the incident light.This energy then passes back into the first medium to form the reflected ray –unless it is absorbed.

Evanescent Wave

Evanescent wave intensity at the interface (z=0)

Exponential decay of intensity (depth)

Exponential decay of intensity (angle)

3D: relative intensity over z and angle

Evanescent wave biosensor

Evanescent wave: removing the cladding near the distal end of the fiberImmuno-probe: covalently attaching antibodies to the coreBinding antigenThe signal: binding the fluorescent-labeled antibodyApplication: clinical diagnosis, pollution monitoring, and process control

Principle

Advantages?

Penetrates a very short distance into the medium.Decays exponentially from the surface. Fluorescence is only excited within a very small volume. Very useful for looking at events near to a surface.Very low background of out-of-focus fluorescence.Decreases the need for washing or separation procedures to divide bound from free ligand, since only the region near the surface of the waveguide is sensed

Why?

Field amplitude of the evanescent wave decreasesexponentially with distance from the wave guideDecreases the need for washing or separation procedures to divide bound from free ligand, since only the region near the surface of the waveguide is sensed

Evanescent Wave

Evanescent wave penetrates less than a wavelength beyond the core - excites the fluorescent molecules Penetration depth (dp):

the distance at which the magnitude of the electric field at the surface decays to its l/e value.

Penetration depth

θ1: internal incident ray angle with the normal to the core/cladding interfacePenetration depth provides a spatial separation between the fluorescent complexes bound to the core and those free in solutionHighly specific antibody binding eventEliminates the need for the washing step

Design: Fluorimeter

CompactUltra-sensitiveDiscrimination between the exciting laser light and the subsequently generated fluorescence, which is much weakerThreshold sensitivity: ability to discriminate a low level of fluorescence.

Optimal probe configuration

Generate and collect fluorescence at the surface of the fiber core.Improves the threshold detection level.

Problems

Removing the cladding causes an abrupt disturbance in the dielectric structure of the optical fiber.Light entering the fiber from the evanescent wave region couples primarily into higher order modes of the unclad fiber.Non-propagating modes in the clad fiber.

Step-etched fiber

nco = 1.458; naq = 1.333; ncl = 1.410Number of modes: V2/2 for step-index fibers

in the sensing region than the clad fiberMore Less

Some of the fluorescent signal will be lost upon entering the clad fiber. (~ 60%)Problem?

Solution

Reduce the core radius of sensing region

Matching radius: 62 µm

Signal versus TRITC-labeled goat-IgG concentration

Results

The unbound material never contributed more than 10% of the signal

small signal decrease upon addition of PBSwashing step was not essential for determination of signal

A 20-fold improvement in the threshold sensitivity3.3 nM (99.5 µm radius) -> 165 pM (52 µm radius)

Three configurationsThe optimal step-etched probe

lost an excessive amount of light where the abrupt decrease in fiber radius occurred. resulted in an increase in bulk fluorescence.

The optimal continuously tapered probedid not achieve a V-number matching radius for almost half its length.

Combination tapered probemaximizes both the power in the evanescent waveand the ability to capture the generated fluorescence.

Fiber probe design

Combination taper

Tapers from the original radius of 100 microns rapidly down to a V-number matching radius of 62 microns.

fluorescence enters the fiber probe and be captured in modes which also propagate in the clad fiber.

Slowly tapers over the remainder of its length to a radius of 37 microns.

permits the power in the evanescent wave to be replenished over its length

Flow Chamber (200 µl)

Bacillus anthracis

Bodily fluids

The ability to detect analytes in biological fluids is critical in using the device in a clinical environment.Accurate determination of the F1 antigen concentration was obtained in serum, plasma, or whole blood Threshold detection level: 5 ng/ml Most clinical immunoassays could be adaptable for use with the fiber optic biosensor.

Effect of bodily fluids on Yersinia pestis F1 antigen fiber optic immunoassay

Measure the fluorescence produced by various concentrations of F1 antigen between 5 and 500 ng/mlExperimentally determined concentration (solid) was compared to the actual F1 concentration (shaded).

Fiber ReuseRegenerate the fiber by eluting the bound material.Requires formulation of a specific elution buffer to facilitate regeneration.The fiber retained significant binding activity over six cycles of use.The response decreased slightly each time the fiber was reused.Primarily due to build up of undissociated antigen, which was photobleached, rather than to a decrease in antibody activity.

Regeneration of the optical fiber’s sensor region

Summary

Field testedPortable multichannel deviceApplications: bedside monitoring in hospitals, effluent monitoring at chemical factories, or screening environmental samples at remediation sites.

ReferencesDevelopment of an evanescent wave fiber optic biosensorAnderson, G.P.; Golden, J.P.; Cao, L.K.; Wijesuriya, D.; Shriver-Lake, L.C.; Ligler, F.S.;Engineering in Medicine and Biology Magazine, IEEEVolume 13, Issue 3, June-July 1994 Page(s):358 - 363 Calibration methods for an evanescent wave fiber optic biosensorGolden, J.P.; Anderson, G.P.; Cao, L.K.; Ligler, F.S.;Engineering in Medicine and Biology Society, 1994. Engineering Advances: New Opportunities for Biomedical Engineers. Proceedings of the 16th Annual International Conference of the IEEE3-6 Nov. 1994 Page(s):822 - 823 vol.2