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Surface-plasmon related ultra sensitive analytical methods and their bio- & nano- applications

Surface-plasmon related ultra sensitive analytical methods and their bio- & nano- applications. Fang Yu, Wolfgang Knoll Max-Planck Institute for Polymer Research, Mainz, Germany. International Congress of Nanotechnology , Nov . 7-10, 2004, San Francisco, USA. Outline. Introduction

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Surface-plasmon related ultra sensitive analytical methods and their bio- & nano- applications

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  1. Surface-plasmon related ultra sensitive analytical methods and their bio- & nano- applications Fang Yu, Wolfgang Knoll Max-Planck Institute for Polymer Research, Mainz, Germany International Congress of Nanotechnology, Nov. 7-10, 2004, San Francisco, USA.

  2. Outline • Introduction • Surface plasmon fluorescence immunoassays • Bridge SPR technology and nano-world • Development of surface plasmon diffraction sensor • Summary

  3. Biosensors additional time and costs for labeling unnatural binding nonlinear signal • e.g. • Surface plasmon resonance (SPR) • Quartz crystal microbalance (QCM) • Reflectometry interference spectroscopy (RIfS) • Surface plasmon diffraction (SPD) • Microcantilever • ... • e.g. • Surface plasmon fluorescence spectroscopy (SPFS) • Total internal reflection fluorescence (TIRF) • ... Labeling Label-free Mass-labels? Beacon tech? insufficient sensitivity non-specific binding complex apparatus

  4. Distinctive features of SPR • Short range phenomenon • Enhanced electromagnetic field • Propagating with high attenuation • p-polarized

  5. Principle of Surface Plasmon Fluorescence Spectroscopy (SPFS) Prism Au water Dipole-to-dipole coupling Surface plasmon Back-coupling Free emission 20 prism 15 100 metal 10 Field-enhancement Factor Fluorescence Yield % dye 5 dielectric 0 0 0 100 z /nm

  6. The set-up photodiode 2 Laser-shutter laser 632.8 nm goniometer prism  lens chopper polarizers attenuator flow cell filter a) PMT b) FOS c) CCD camera PC shutter controller motor- steering photon- counter lock-in amplifier

  7. Surface matrix for biosensing • Good chemistry (for NSB, activation, regeneration…) • Lateral control of ligand density • Compatible with the physics of the biosensor Ideally… • 2D (SAM, lipid bilayer…) • 2.5D (Layer-by-layer, nano-particle, nano-capsule, nano-wire modified surface, porous or roughened surface…) • 3D (brush type polymer, hydrogel network, plasma- or electro-polymerized matrix…) Dimension…

  8. Interfacial design of sensing matrix Au water : antibody 2D (e.g. layer-by-layer assembly) : fluorophore : streptavidin 3D (e.g. dextran matrix) : SAM : dextran

  9. LbL to clarify metal-induced quenching Alternating biotin-IgG and SA, decorate certain layer by Alexa fluor labeled SA IJ* : Ib* = 34

  10. Surface preparation for dextran matrix (2) (3) (1) (3) (2) ~8 ng mm-2 (1)

  11. Limit of detection (LOD) evaluation under mass-transport limited binding condition Baseline deviation

  12. Correlation between SPR and fluorescence LOD at atto-molar level Translate the LOD level to molecular surface concentration ~10 molecules/(mm2*min) Yu, F., Persson, B., Loefas, S., Knoll, W. JACS, 126, 8902 -8903, 2004.

  13. D=kT/6 a Prostate-specific antigen (PSA) sandwich assay slopekm= 0.98(D/h)2/3(v/bx)1/3 -2/3

  14. LOD of PSA assay without plasma NSB Yu, F., Persson, B., Loefas, S., Knoll, W. ANALYTICAL CHEMISTRY, in press.

  15. Streptavidin-latex bead in SPR sensing SA doping ratio: ~300 SA per bead 125 nm Utilities: 1, Signal amplification 2, Introduce surface scattering 3, Being functional matrix itself (2.5 D)

  16. FOS (Fiber optic spectrometer) SA-Lx Biotin SAM (1:9) Au Surface plasmon enhanced light-scattering 633 nm laser

  17. (1) (2) (3) (4) (5) Coverage dependent scattering

  18. One step SPR detection of 15mer oligonucleotide by latex-amplification

  19. DNA conjugated core/shell QDs Core/shell QDs supplied by Q-Dot Corp. : high stability in PBS before and after conjugation wavelength 565nm (green), 585nm (yellow), 605nm (orange) and 655nm (red) are all excitable with 543nm (green laser) 5’-biotinylated target DNA

  20. P1 P1 P1 P2 P2 P2 P3 P3 P1+ P1+ P2 P2 Color Multiplexed hybridization detection test Excitation-Filter (543nm) +QD565-T2 (MM0 for P2) +QD655-T1 (MM0 for P1) Microarray image from SPM P2 P1 Robelek, R., Niu, L., Schmid, E. L., Knoll, W. ANALYTICAL CHEMISTRY, in press

  21. Principle of SPDS -2 -1 Diffraction orders (m) 0 1 2 Functional area Nonfunctional area Au Dielectric grating nd, the grating amplitude 

  22. TIR diffraction vs. SPR diffraction -2 -1 0 1 2 -2 -1 0  1  2 glass Au  Polystyrene pattern TIR mode ATR/SPR mode

  23. Diffraction patterns Surface plasmon microscopy images Diffraction photographs

  24. Micro-contact printing for SAM patterning 4 Photoresist PDMS Si 1 5 Photoresist pattern PDMS Si Au 2 6 PDMS Si Au 3 Functional SAM 7 Nonfunctional SAM PDMS Au

  25. Quadratic property of the diffraction signal Anti-biotin antibody Biotin SAM Yu, F., Tian, S., Yao, D., Knoll, W. ANALYTICAL CHEMISTRY, 76, 3530 -3535, 2004.

  26. Self-referencing property of the diffraction sensor - a temperature variation test Yu, F., Knoll, W. ANALYTICAL CHEMISTRY, 76, 1971-1975, 2004.

  27. 500 nM Fab-biotin • 50 nM hCG • 50 nM hCG in 1 mg/mL BSA • 1 mg/mL BSA hCG SPDS for label-free detection of human chorionic gonadotropin (hCG) Fab SA SAM

  28. SPDS for oligonucleotide detection 1, surface preparation target DNA probe DNA SA biotin SAM nonfunctional functional

  29. SPDS for oligonucleotide detection 2, kinetic analysis HE: Hybridization efficiency Yu, F., Yao, D., Knoll, W. NUCLEIC ACIDS RESEARCH, 32, e75, 2004.

  30. SPDS for oligonucleotide detection 3, adsorption isotherm analysis KD0 KD1

  31. Summary • Ultra-sensitive SPFS immunoassay is established with the aid of three-dimensionally extended matrix • Initial attempts of SPR based nano-sensing • SPDS is developed for label-free analysis of protein interactions and oligonucleotide hybridizations

  32. Acknowledgements Neal Armstrong (University of Arizona) Akira Baba (University of Texas at Houston ) Shengjun Tian (MPIP) Lau King Hang Aaron (IMRE, Singapore) (for helps in the diffraction work) Björn Persson (Biacore) Stefan Löfås(Biacore) Renate Sekul (Graffinity) Holger Ottleben (Graffinity) (for collaborations) Danica Christensen (MPIP) (for the LBL work) Pierre Thiébaud (MPIP) Darick Ding (MPIP) (for the set-up engineering ) Danfeng Yao (MPIP) Thomas Neumann (Graffinity) Eva - Kathrin Sinner (MPI biochemistry) Peter E. Nielsen (Panum Institute, Denmark) Keiko Tawa (AIST Osaka) Rudi Robelek (IMRE, Singapore) Lifang Niu (IMRE, Singapore) (for the DNA/QDs part)

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