jussi hiltunen photonics days 2017, summer school
TRANSCRIPT
Raman measurements
Jussi HiltunenPhotonics Days 2017, Summer School
Outline
• Introduction to Raman scattering-Principles-Instruments-Surface enhanced Raman spectrosopcy (SERS)
• Microsystem integration• Analyte size effects
Raman spectroscopy§ Is a form of vibrational spectroscopy, measures inelastic light scattering§ Complementary to IR spectroscopy, different selection rules§ Most of molecular vibrations can be found in IR and Raman spectra
sample
sample
IR
Raman
Each material hasown finger print
Origin of Raman scattering
• Dipole moment is a measure of the separation of positive and negative electrical charges within a system• Electric polarizability is the relative tendency of a charge distribution displacement by an external electric• Coupling of light with dipole moment change induces absoprtion -> IR spectroscopy• Coupling of light with the change in polarizability induces Raman scattering
0( ) ( ) ( )
0 0
Mathematical formulation
Input field Induced polarization
Displacement dependent polarizabilityTime-dependent displacement
Coupling of input and vibrational frequences
Original frequency Coupled frequencies
Energy state diagram
• Rayleigh scattering is elastic process without energy transfer• In Stokes shift, the final state is higher in energy than the initial state.• In anti-Stokes shift, the final state is lower in energy than the final state.
Raman instrument
Laser
Laser filterSample
Emission filter
Spectrometer
• Raman instrument is essentially a spectrometer with a narrow band light source• System can be high-end laboratory instrument or hand-held device – and anything between
RingC-H
C-S
Benzenethiol spectrum
Wavenumber (1/cm)
Inte
nsity
Wikipedia
BWTekBWTekBaySpecTimeGate
Surface Enhanced Raman Scattering (SERS)
by Sanna Uusitalo
Surface plasmon polariton
by Sanna Uusitalo
• Strong field localization can occur due to the couplingof charge motion in the metal (surface plasmon) andelectromagnetic waves in the air or dielectric(polariton).
• Field effects can induce high enhancement of Ramanscattering 10^4 and 10^8
• Enhancement is localized near the surface
Nanoparticles
Nanostructured surfaces
UV-imprinted surface enhancedRaman spectroscopy substrates
Cut-out 96-well plate size sheets withSERS patterns
• A plastic SERS sensor was replicated by roll method• The plastic web was die-cut into sheets with 1cm*1cm
SERS patterned squares• Cut-out SERS sheets were coated with gold layer
www.photosens.eu
SERS sensor with integrated microfluidics
SERS layer
Fluidics layer (double sided tape)
Lid (polyolefin)
SERS enhancing structures were integrated to study the formation of signal in embedded structures1) Dynamics2) Background level
On-chip detection of Rhodamine 6G model analyte
• 1mM R6G model analyte sample
• The Raman spectrums recorded ontop of the SERS area and on thesmooth gold area for reference
→ The detected signal from theSERS area is surface enhanced
1100 1200 1300 1400 1500 1600 1700 1800
0
10000
20000
1187
1313
1361
1511
1600
1181
1309
1367
1513
Ram
anin
tens
ityRaman shift [1/cm]
1mM R6G on SERS Reference R6G on gold
Parameters used with Raman microscope:- 785nm laser- 20X objective- Integration time 15s- 2*3 point Image mapping- No signal accumulation
Continuous flow of 0.5mM R6G on-chip• Fluids were transferred from a sample
vial into the chip by underpressure• Water was first loaded into the chip
for base reference• R6G sample followed and the signal
rise was detected for flow velocities of25µl/min to 1000µl/min
• Trial was conducted under afluorescence and a Raman microscope
Continuous flow of 0.5mM R6G on-chip• Signal response for the convective flow of R6G molecules detected by fluorescence
microscopy• Diffusion affected flow at the sensor surface detected by SERS
0 2 4 6 8 10
0.0
0.2
0.4
0.6
0.8
1.0
Nor
mal
ised
sign
alin
tens
ity
Time (min)
25µl/min 30µl/min 50µl/min 250µl/min 500µl/min 1000µl/min
0 5 10 15 20
0.0
0.2
0.4
0.6
0.8
1.0
Nor
mal
ised
sign
alin
tens
ity
Time [min]
25µl/min 30µl/min 50µl/min 250µl/min 500µl/min 1000µl/min
SERS resultsFluorescence results
Effect of the polymer lid to the Raman spectrums• Polymer materials are Raman active
• Polyolefin lid has Raman peaks, no fluorescencebackground with 785nm
• The distortion of the R6G peaks can be removed bysubtracting the reference spectrum from the finalresults
Parameters used withRaman microscope:- 785nm laser- 20X objective- Integration time 30s- 2*3 point Image mapping- No signal accumulation
Background substracted 10µM and 100µM R6G spectraRaw spectra
Analyte size effects in SERS
Molecules
• Typically microbe size is in the order of µm while plasmonic features are below 1 µm-> Plasmon field can probe only the part of the ”analyte”
Sample preparation for size effect studies
• Listeria bacteria cells were cultured and immuno-magentic bead-separated (IMS) prior dispensing on the SERS substrate• Au nanoparticles were subsequently dispensed• As a result hybrid nanoparticle-nanosurface configuration was formed
Detectedlines
Raman shift(cm-1) Assignments
631 627/620 Phenylalanine (skeletal)
737 732
glycosidic ring mode of D-glucoseamine (NAG), adenine or
CH2 rocking968 955 C-N stretching
1142 1134/1130C-N and C-C stretch
(carbohydrates)1271 1230-1295 Amide III
13391334/1339/133
8
Deformation CH/Amide III/signature of adenosine;
monophosphate and guanosinemonophosphate, aromaticamino acids tyrosine and
tryptophan1374 1371 DNA
1397 1392/1398Symmetric deformation of CH3
groups1450 1453 CH2 deformation (lipids)
Raman signal intensity with different configurations
• Three configurations were tested1) Listeria on corrugated SERS substrate2) Listeria surrounded with Au nanoparticles, but without substrate enhancement3) Listeria surrounded with Au nanoparticles and located on SERS substrate
• Highest signal was obtained from the hybrid structure
AcknowledgementsSanna Uusitalo, PhD candidate
Thank you for your attention!