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Alexandre FRAN Ç OIS , Sivashankar KRISHNAMOORTHY and Michael HIMMELHAUS

Novel detection scheme for optical biosensing using whispering gallery modes in clusters of dielectric particles. Alexandre FRAN Ç OIS , Sivashankar KRISHNAMOORTHY and Michael HIMMELHAUS. Bio-Nanotechnology Research Project – Frontier Research Dept.

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Alexandre FRAN Ç OIS , Sivashankar KRISHNAMOORTHY and Michael HIMMELHAUS

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  1. Novel detection scheme for optical biosensing using whispering gallery modes in clusters of dielectric particles Alexandre FRANÇOIS, Sivashankar KRISHNAMOORTHY and Michael HIMMELHAUS Bio-Nanotechnology Research Project – Frontier Research Dept. FUJIREBIO, INC., 51 Komiya-cho, Hachioji-shi 192-0031 JAPAN

  2. What are Whispering Gallery Modes? When light is trapped in a micro sphere by Total Internal Reflection (TIR), it might circulate within the sphere along its circumference. These modes are so-called Whispering Gallery Modes (WGM).

  3. Whispering Gallery modes for sensing Any modification of the micro spheres optical properties (adsorption of molecules at its surface) results in modification of the WGM resonance frequency. WGM frequencies are influenced by both the micro sphere shape (radius, sphericity) and optical properties (index of refraction, coefficient of absorption and scattering). Courtesy of J.U. Nockel (http://www.uoregon.edu/~noeckel/microlasers.php)

  4. Why is this interesting for biosensing ? • label-free • highly sensitive (less than femtomoles) • non-destructive to the sample • highly selective (through surface functionalization of the micro sphere surface) • applicable to various targets • fast and simple (wireless) excitation & detection

  5. Microresonator High Q WGM sensor Low Q WGM sensor • External excitation via optical coupling with an eroded optical fiber. • Require a tedious tuning of the gap between the microsphere and the optical fiber (nm scale). • Microsphere diameter ranging from 60 to 300 mm which hamper the prospect of integration of many microsphere within the same microfluidic flow cell. • Internal excitation with fluorescent dye. • Allow a reduction in size to increase the WGM wavelength shift. • Many resonance peaks available Radius of the microsphere can be calculated from the WGM spectrum. Arnold et al. OPTICS LETTERS (2003) 28, 4, 272-274 Guo et al. J. Phys D: Appl. Phys. (2006) 39, 5133-5136

  6. Experimental setup Flow cell containing beads Outlet Inlet CW Laser excitation l=442 nm 15 mm Inverted microscope Detector: PMT or CCD camera Optical fiber Monochromator

  7. WGM spectrum 2nd order WGM TEm+1 TMm+1 TEm TMm 1st order WGM

  8. Detection scheme of WGM for optical bio sensing • Measurements of the WGM of single beads as reference spectrum prior to any modification of the bead surface. • Adsorption of the bio molecule on the bead surface. • Measurements of the WGM after Adsorption of the bio molecule. A wavelength shift of the WGM indicates whether the bio molecule has been successfully adsorbed. • Single bead position has to be precisely recorded. • Single beads cannot be differentiated by means of their WGM spectra.

  9. New detection scheme using clusters of beads instead of single bead All clusters exhibit a particular spectrum that can be considered as a fingerprint.

  10. Wavelength shift of WGM of a single bead vs. cluster of beads after adsorption of polyelectrolyte PAH polyelectrolyte coating (~ 3nm) PSS PS Bead Successive deposition of polyelectrolyte results in an increase of the bead radius. Wavelength shift of the WGM spectrum towards higher wavelength for both single bead and cluster of three beads. (1) Reference spectrum in MP water. (2) 1st PE coating. (3) 2nd PE coating. (4) 3rd PE coating.

  11. Autocorrelation function as a tool to identify clusters Autocorrelation between two spectrum from the same single microsphere Autocorrelation between two spectrum from the same cluster of 3 microspheres Autocorrelation between two different single microsphere Autocorrelation between two different clusters of 3 microspheres

  12. Is the WGM wavelength shift depending on the number of beads in the cluster? NO !!!! Whatever is the number of beads present in the considered cluster, the WGM wavelength shift is the same.

  13. R R+DR Relation between WGM wavelength shift and increase in bead’s radius DR = deposited thickness R = bead radius (5mm) Dl = WGM wavelength shift l = WGM wavelength nS= bead refractive index (1.59) nL= PE refractive index (1.47) Measurements are in good agreement with the expected thickness of PE coating [1] M. Lösche et al., Macromolecules (1998) 31, 8893-8906 [2] Caruso et al., Science (1998) 282, 1111-1114

  14. Detection of a protein with WGM Dlmeasured = 0.15 ± 0.01 nm sP-1calculated = 4.5×10-13 cm2 sP-1BSA = 3.5×10-3 cm2 [2] Which is consistent with no more than one monolayer. [1] Projected area : ns : bead refractive index (1.59) nm : surrounding medium refractive index (1.33) aex :excess polarizability (4p×3.85×10-21 cm3) [1] [1] Arnold et al., OPTICS LETTERS (2003) 28, 4, 272-274 [2] Ferrer et al., Biophysical Journal (2001) 80, 2422–2430

  15. Detection of a protein with WGM Comparison of BSA adsorption on a single microsphere with SPR Kinetic WGM spectrum of BSA adsorption on a single microsphere Detection of BSA adsorption by WGM gives similar results compared with SPR

  16. Comparison with state of art WGM sensor • The molar resolution increases as the microspheres radius decreases. • Our sensor is 100 times more sensitive compared to the state of art WGM sensor even with low detection sensitivity and “poor” quality factor. [1] Arnold et al., OPTICS LETTERS (2003) 28, 4, 272-274 [2] Ilchenko et al., Proc. of SPIE (2007) 6452, 64520U-1

  17. Conclusion Detection of adsorbed molecule using WGM has been done in vitro using single 10 mm dye doped polystyrene spheres. New detection scheme has been found based on clusters of micro spheres instead of single spheres. • It has been shown that clusters exhibit a particular WGM spectrum that can be considered as a fingerprint. • After adsorption of a molecule onto the cluster surface, a WGM wavelength shift occurs that does not depend on the number of microspheres constituting in the cluster. • All calculations following the adsorption of either polyelectrolyte and BSA have confirmed that the sensor based on WGM excitation in clusters is reliable and has the highest molar sensitivity reported so far. • This detection scheme based on clusters of microspheres enables the integration of a large number of sensor inside a microfluidic flow cell

  18. ACKNOLEDGMENT Dr. Yoshihiro ASHIHARA Ms. Tomoko FUJII

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