1 dna detection by differential perturbation of two ...sphere s1 is detected by an increase of the...
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DNA Detection by Differential Perturbation of two Microsphere CavitiesFrank Vollmer1, Stephen Arnold2, Dieter Braun1, Iwao Teraoka2, Albert Libchaber1
1Center for Studies in Physics and Biology, The Rockefeller University, New York, NY 10021email: [email protected]
2Microparticle Photophysics Lab, Polytechnic University, Brooklyn, NY 11201www.poly.edu/microparticle
We report the detection of unlabeled DNA oligonucleotides on microsphereprobes from a response of their photonic resonance modes. Narrow linewidthresonant modes (called whispering-gallery-modes, WGMs) occur from a lightray orbitting inside the microsphere due to total internal reflection. The reso-nance wavelength increases when dielectric material such as nucleic acid isadded on the microparticles surface. Each microsphere is modified with anoligonucleotide probe of interest. The kinetics of an hybridization event isdetected with millisecond time resolution from an increase of a sphere spe-cific resonance wavelength. We demonstrate that the narrow, microspherespecific resonances are separated by several linewidths which allows multi-plexing of signals from two (or more) different microparticle probes coupledto the same waveguide. We show that the differential signal of two micro-spheres can be used to identify a single nucleotide mismatch in a label-free11-mer oligonucleotide. This approach to hybridization measurements is par-ticularly sensitive, with the typical differential signal for a single nucleotidemismatch at 54:1.
λ ... wavelength (1.3 µm)δλ ... wavelength shiftαex ... excess polarizability of the bound proteinσs ... surface density of the bound proteinn1, n2 ... refractive indices of the sphere and the buffer solution, respectivelyR ... orbital radiusε0 ... vacuum permittivity
FV is supported by a fellowship of the Boehringer Ingelheim Fonds, DB by a Fellowship of the Deutsche Fors-chungsgemeinschaft, research at the Polytechnic is supported by a National Science Foundation grant.
microsphere
eroded optical fiber
A light ray can be confined inside a dielectric sphere due to totalinternal reflection at the sphere surface. The long confinement time(high Q) allows the light to circumnavigate the sphere for manyorbits. If used for biosensing, such an optical resonance (WGM) enables thelight to interact with the same analyte molecule for several thousandtimes. The resonance thus improves the detection limit by orders ofmagnitude as compared to existing single pass techniques. The ulti-mate detection limit of an optical cavity has been estimated to be onthe order of a single, unlabeled macromolecule.
Resonances in glass microspheres can beexcited by the light transmitted through anoptical fiber. Coupling of light between fiber and sphereoccurs only for specific resonance wave-lengths. Experimentally, a resonance is detected as adip in the spectrum of the light intensitytransmitted through the fiber-sphere system.Detection of biomolecules is possible due tothe evanescent field which extends from themicrosphere surface. Dielectric material suchas DNA and protein molecules polarize whenentering the evanescent field of the micro-sphere. This perturbation of the optical cavityleads to a red shift of a given resonancewavelength.
(A) Light from a tunablelaserdiode L (1.3 µm) istransmitted through a singlemode optical fiber F. Two sil-ica microspheres S1 and S2are evanescently coupled tothe fiber. A photodetector Precords the intensity at theother fiber end. Optical reso-nances from each sphere areidentified as Lorentzian dipsin the transmission spectrum.A hybridization event of alabel-free oligonucleotide onsphere S1 is detected by anincrease of the S1-specificresonance wavelength.
(B) Micrograph of twospheres coupled to the sameoptical fiber running horizon-tally through the center of theimage. The image shows res-onances of light orbittinginside each sphere.
Sphere Multiplexing. (A) Transmis-sion spectrum for one (dotted line) andtwo (solid line) spheres coupled to thesame optical fiber. Both spheres aremodified with 27-mer oligonucle-otides.
Shift of Resonances. (B) Shows thetime trace of the two resonance posi-tions from S1 and S2. The arrows indi-cate when the two complementaryDNA oligonucleotides were injectedinto the sample solution. Hybridizationsaturates within minutes and the reso-nance wavelength of the correspondingsphere increased about .038 nm each.
Single nucleotide mismatch discrim-ination. (A) Time traces of resonancewavelengths in two spheres S1 and S2.S1 was modified with a 11-mer oligo-nucleotide (CTATCTCAGTC). Theoligonucleotide immobilized on S2differed by a single nucleotide(CTATATCAGTC). The arrow indi-cates when the oligonucleotide com-plementary to the sequenceimmobilized on sphere S1 wasinjected. In equilibrium, the wave-length shift for the perfect matchsequence was approximately ten timesas large as the shift for the mismatchsequence. (B) The difference signalallows one to identify a single nucle-otide mismatch with a high signal-to-noise of 54.
References:“Protein detection by optical shift of a resonanc microcavity”, Applied Physics Letters 80 (21), 2002,4057-4059“Shift of Whispering Gallery Modes in Microspheres by Protein adsorption”, Optics Letters 28 (4),2003, 272-274
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