1. SERS-based Biosensors
James Krier, Lalitha Muthusubramaniam
Kevin Wang, Douglas Detert
Final Presentation
EE235: Nanofabrication
May 12, 2009

2. Overview Technology Landscape: Optical techniques for biosensing
Surfaced-enhanced Raman scattering (SERS)
Technical background
SERS-based biosensors
Financial and market considerations of SERS

3. Vast Technology Landscape
Diverse Applications

4. Total internal reflectance fluorescence (TIRF) biosensor
Evanescent wave
TIR - propagating light encounters a boundary to a medium of lower refractive index, at angle greater than “critical angle”
reflected light generates a highly restricted electromagnetic field adjacent to the interface in the lower ref. index medium
ev. Wave is identical in frequency to the incident light, and decays exponentially in intensity with distance from the interface (high S/N)
possible to do single molecule fluoresence

5. Typical TIRF Sensogram
Epifluorescence
Advantages
High Signal to noise ratio (very little secondary emission from bulk solution)
Highly robust, low cost, portable
Drawbacks
Need for labels
High cross-reactivity (hence not easy to multiplex)

6. Molecularly Imprinted Polymers as Optical Sensors
Schematic representation of molecular imprinting
This is a process where functional and cross-linking monomers are copolymerized in the presence of a target analyte (the imprint molecule), which acts as a molecular template. The functional monomers initially form a complex with the imprint molecule, and following polymerization, their functional groups are held in position by the highly cross-linked polymeric structure. Subsequent removal of the imprint molecule reveals binding sites that are complementary in size and shape to the analyte. In that way, a molecular memory is introduced into the polymer, which is now capable of selectively rebinding the analyte
Despite the conceptual similarities between MIPs and antibodies, they display very different types of recognition properties. The most significant difference is that MIPs contain a heterogeneous distribution of binding sites, whereas monoclonal antibodies are homogeneous and have a single type of binding site. The concentration-dependant binding properties can be exploited at low analyte concentrations.
Distribution of binding affinities in MIP vs. Ab
Chemical Reviews, Chem. Rev., (2000)

7. 3 methods to monitor binding in MIPs
Direct monitoring of analyte in solution; Incorporation of spectroscopically responsive monomers into the matrix;Competition assays using labeled ligands
Challenges in developing fluorescent and UV-visible monitored MIP sesnors:
(1) low selectivities and high cross-reactivities of MIPs; (2) lack of an inherent signaling mechanism in MIPs; (3) poor optical and material properties of MIPs.
Polymer International, Vol 56( (4), pp

8. Reflectometric interference spectroscopy (RIFS)
The reflected beams superimpose and change optical thickness of the transducer by binding events onto the surface. Shift in characteristic interference spectrum is transformed into a signal curve.
J. Immunological Methods Vol 292, Issues 1-2, September 2004, pp.35-42

9. Reflectometric interference spectroscopy (RIFS)
Protein concentration determined spectrophotometrically and active antibody concentration determined by biosensor and ELISA for 9 sequentially eluted fractions.
J. Immunological Methods Vol 292, Issues 1-2, September 2004, pp.35-42

10. The SERS Solution
Adsorption
Excitation
Detection

11. Raman Spectroscopy C.V. Raman
Adapted from

12. Provides rich info. about structural data!
Raman Spectroscopy
Selection rules
Based on symmetry elements of polarizability tensor
Vibrational fingerprint
Comprised of narrow spectral features
Robust mechanism
Not subject to photobleaching
Weak Signal
Compared to Rayleigh scattering / fluorescence
Provides rich info. about structural data!
adenine
cytosine
guanine
thymine
uracil
Gelder, et al., J. Raman Spectrosc., (2007)
A. Campion et al., Chem. Soc. Rev., (1998)

13. Surface-Enhanced Raman Scattering
1928
C.V. Raman discovers “Raman Effect” of inelastic scattering
1974
Discovery of enhanced Raman signals ( ) from molecules adsorbed on roughed Ag surfaces.
Mechanism is attributed to enhanced surface area for adsorption.
1977
Debate begins over the exact mechanism of signal enhancement.
M. Fleischmann, et al., Chem. Phys. Lett., (1974)
D.L. Jeanmaire, R.P. Van Duyne, J. Electroanal. Chem., 84 1 (1977)
M.G. Albrecht, J. A. Creighton, J. Am. Chem. Soc., (1977)
S. Schultz, et al., Surface Science, (1981)
M. Moskovits, , Reviews of Modern Physics, 57 3 (1985)
K. Kneipp, et al., Chem. Rev., (1999)

14. Away from plasmon resonance
SERS Enhancement
Chemical Enhancement
Based on metal-molecule charge- transfer effects
Electromagnetic enhancement
Coupled to surface plasmon excitation of metal nanostructures
Tunable resonances: Shape- and Size-effects
Away from plasmon resonance
At plasmon resonance
A.J. Haes, et al., Anal. Bioannal. Chem., (2004)
S. A. Maier, et al., Adv. Mater., (2001)

15. SERS Enhancement
Plasmon resonance leads to local field enhancement near the surface
Adsorbed molecules see increased field
Raman signal enhancement (up to 1015)
Depends on local geometry of adsorption site
Enhancing SERS substrates nm
K. Kneipp, et al., Chem. Rev., (1999)
J. Aizpurua, et al., Phys. Rev. Lett., (2003)

16. The SERS Advantage Molecular fingerprinting
Unique vibrational spectra distinguishes molecules
Tagless biosensing
Fluorescent dyes are not needed
Multiplexed sensing
Plasmon resonances allow for sensor tunability
In vivo applicability
Near-IR excitation and biocompatability allow
Femtomolar and beyond
Single molecule spectroscopy is possible
1500 cm-1
1532cm-1
1600cm-1
1635cm-1
S.M. Nie, et al., Science, (1997)

17. Single Molecule Detection
PRL 78, 1667 (1997)

18. TERS
nanowerk.com

19. TERS
Faraday Discuss., 132, 9 (2006)

20. TOPOGRAPHY + SPECTROSCOPY
PRL 100, (2008)

21. In-vivo glucose sensing
Faraday Discuss., 132, 9 (2006)

22. Other Options
PRL 62, 2535 (1989).

23. More Moerner et al.
Nature 402, 491 (2000).

24. stanford.edu/group/moerner/sms_movies.html

25. NSOM
JPC 100, (1996)

26. SERS Market Consumables $50 to $750 per analysis
$1 million market annually
Instrumentation
$10,000 - $180,000
Image source: (New York Talk Exchange)
Numbers:
Image source: (New York Talk Exchange)
Numbers:

27. SERS Companies Renishaw Real Time Analyzers Bruker Optics
D3 Technologies (Mesophotonics)
Oxonica
Renishaw
Real Time Analyzers
Source:

28. SERS Vials Real Time Analyzers Sol-gel of Au or Ag nanoparticles
106 signal enhancement
Source: RTA

29. Portable Raman Real Time Analyzers RamanID DeltaNu Inspector Raman
Diesel Fuel Spectrum

30. SPR Companies Biacore (GE) Biosensing Instrument FujiFilms
GWC Technologies
Ibis
Sensiq

31. SPR Analyzer Biosensing Instrument BI- 2000 Cost: $39k
Liquid/Gas Detection
10-4 degree sensitivity

32. Cost Comparison Method Equipment Consumables SERS
Spectrometer, $10kHe/Ne Laser, $760Optics, $100Microflow Cell, $300Total = $11.1k
Au Nanoparticles ($3/mL)
TERS (AFM+ SERS)
AFM ($90k - $150k)Total = $111k - $161k
AFM tips ($10)
SPR
Full Setup, $39k - $60k
NSOM
Full Setup, $100k - $250k
NSOM tip ($100)

33. Conclusion: SERS
Even simple (diatomic) molecules can have complex and reproducible vibrational fingerprints
The most practical option for sensing near the single-molecule level for a variety of analytes in solution or air, lending to an array of applications ranging from trace gas detection to automated protein identification
Easy to couple with other supplementary techniques (e.g., AFM)
Provides an economically feasible sensing mechanism for portable devices in atmospheric conditions