Transcript Slide 1

SERS Biosensor for Endocrine
Disruption Biomarker: Vitellogenin
Sachin Kumar Srivastava
Mentor: Prof. Ibrahim Abdulhalim
Department of Electro-optic Engineering, &
Ilse Katz Institute for Nanoscale Science and Technology,
Ben-Gurion University of the Negev, Be’er Sheva, Israel
Outline
 Introduction
 Raman-/Surface Enhanced Raman- Spectroscopy (SERS)
 Biosensor
 Endocrine Disruption Compounds
 Nano-Sculptured Thin Films (n-STFs): Optimization of performance
 Fabrication : Steps involved in developing the sensor
 Results: Sensing and other characterizations
 Summary
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Raman Spectroscopy
Indian Physicist: C. V. RAMAN
"A small fraction of sunlight passing through a transparent medium is
scattered in all directions, with a different wavelength than the incident
radiation. These shifts depend upon the chemical structure of the molecules.“
"A new radiation", Indian J. Phys.,2(1928)387.
Nobel prize in Physics : 1930
Raman Scattering
Rayleigh Scattering
Lowest
Electronic
State
v3
v2
v1
v0
Ground State
Anti-Stokes
Stokes
Virtual levels
Rayleigh Scattering: Elastic
Raman Scattering: Inelastic
1 in 107 photons is scattered
inelastically
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‘Fingerprint’ of molecular bonds and crystalline structure
Very low cross-section: Applications in real life are hindered
Steps of Raman development
• Laser applications – Dramatically improved power of excitation and Raman signal
• SERS effect – Enhanced intensities up to factors of 108-9
• Raman microscope – Decreased probe volume (light spot diameter below 1 μm)
What is SERS?
Large local field enhancement on rough metallic surfaces due to plasmonic and charge
transfer enhancement mechanisms
p   Elocal
Incident light
Enhanced Raman Scattering
Plasmonic Nanostructures
Adsorbed molecules
SERS Substrate
Nanosculptured Thin Films (nSTFs)
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Biosensor
Sensor
A device having the role of converting a change in one physical parameter into
a change in magnitude of a second different parameter which can be measured
more conveniently and perhaps more accurately.
Bio-sensor
A biosensor is a self-contained integrated device that is capable of providing
specific quantitative or semi-quantative analytical information using a biological
recognition element which is in direct soatial contact with a transduction element
(IUPAC, 1996)
Any device that uses specific biochemical reactions to detect chemical compounds
in biological samples.
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Components of a Biosensor
/ Optical
Molecule
imprinted gel
matrix
Chemical
interaction
SPR/SERS etc
Detector
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Basic Characteristics of a Biosensor
1. SENSITIVITY: Response per unit analyte numerical value.
2. DETECTION ACCURACY: How accurate the measurement is?
3. LIMIT OF DETECTION: What is the minimum numerical value
of the parameter which can be sensed?
4. SELECTIVITY: Interference from other factors must be
minimized for obtaining correct results
nm1<nm2
5. RESPONSE TIME: Time necessary for having about 95% of the response. Wavelength
6. REUSABILITY: Can the sensor be used again and again?
7. SHELF LIFE: What happens to the performance when kept for long time
8. SENSING VOLUME: Volume of analyte required for sensing
9. RANGE OF OPERATION: What is the working range of the sensor
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Endocrine System
General
Functions of
Hormones
http://www.cea1.com/anatomy-sistems/function-ofthymus-in-endocrine-system/attachment/functionof-thymus-in-endocrine-system/
Bond 1979, Biology of Fishes
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Endocrine Disruptors
Metals
(Pb, As, Hg, Ni, Cd)
Personal Care
Products
(Phthalates,
Benzaldehyde)
Pesticides,
Herbicides etc.
Growth inhibition
Diabetes
Industrial
Chemicals
Suspected
Endocrine
Disruptors
(DDT )
(Bisphenol A)
Synthetic and
Natural Hormones
(Phytoestrogens)
Pharmaceutical
Drugs
(Corticosteroids)
Vg : Egg yolk protein in aquatic
females, excreted during sex
Elevated amounts: A sign of
estrogenic contamination of aquatic
environment.
 Representative of a kind of
pollution in water/aquatic species
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nSTF
Fabrication
GLAD Technique
SEM Images
(20, 30, 40, 60 % porosity)
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Optimization of Performance
SERS from 4-Aminothiophenol (4-ATP) adsorbed at the n-STF
 Highest SERS signal due to
plasmons coupling
 Mutual interaction between
the EM fields of the localized
surface plasmons (LSPs) at the
neighbouring nanorods
 Effective
electromagnetic
field of the LSPs in the voids
 Field in voids decreases with
increase in porosity
 The number of adsorbed
molecules increases, however.
Maximum Enhancement @ 30% porosity
Trade off between the
enhancement due to rods and
voids
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Sensor Chip Development
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Experimental Setup
From Laser
SERS
Probe
To Spectrometer
Anti-Vg Antibody
Vitellogenin
BSA
Extended view of SERS Probe
Sensor Chip
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Comparison of SERS Spectra@ Sensor vs 4-ATP
Small range effect (~ 2-3 nm)
Negligible effect of further added molecules
Dieringer and van Duyne et al., Faradey Discuss., 2006, 132, 9-26
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Sensor Response for Varying Vg Concentration
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Control Experiments
 Confirmation for the specificity of the sensor to vitellogenin.
 Another similar protein called Fetuin was used for study.
Reusability and Repeatability
 A number of sensor chips were prepared and experiments were performed
over a period of three months at different days, times and ambient conditions
 The sensor surface was regenerated by immersing in 50mM glycine buffer for
2 minutes in between two sensing events.
 Each chip was used at least three times for full sample concentration range to
ensure the repeatability and reusability.
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Sensor Response Curve
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Summary
 A highly sensitive, fast Vg sensor chip utilizing SERS has been fabricated
 SERS can be used for sensing of bigger molecules in non-direct mode of sensing
 Vitellogenin concentrations as small as 5pg/ml can be detected
 The sensor is specific to Vg only.
 The repeatability, reusability and shelf life of the sensor are reasonably good
For more detailed discussion of the presented results, please see
SERS Biosensor Using Metallic Nano-Sculptured Thin Films for the
Detection of Endocrine Disrupting Compound Biomarker Vitellogenin,
SMALL (Wiley), 2014. DOI: 10.1002/smll.201303218
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Other Contributors/Co-authors
Dr. Atef Shalabney,
Laboratoire des Nanostructures, ISIS - Institut de Science et d'Ingénierie
Supramoléculaires,Université de Strasbourg, 8 allée Gaspard Monge 67000Strasbourg, Cedex
Dr. Isam Khalaila,
The Avram and Stella Goldstein-Goren Department of Biotechnology Engineering,
Ben Gurion University, Beer Sheva-84105, Israel
Dr. Christoph Grüner and Prof. Bernd Rauschenbach
Leibniz Institute of Surface Modification, Permoserstrasse 15, 04318 Leipzig and
University Leipzig, Institute for Experimental Physics II, Linnéstr.5, 04307 Leipzig,
Germany
Prof. Ibrahim Abdulhalim
Department of Electro optic Engineering and Ilse Katz Institute for Nanoscale
Sciences and Technology, Ben Gurion University of the Negev, Beer Sheva 84105,
Israel &
School of Materials Science and Engineering, Nanyang Technological University,
637722, Singapore
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Acknowledgements
1. Council of Higher Education of the Government of the State of Israel
for PBC post-doctoral fellowship
2. Singapore National Research Foundation (CREATE programme)
3. German Excellence
Gemeinschaft (DFG).
Initiative
of
the
Deutsche
Forschungs
Thank You
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Dipole moment in a molecule
p E
α0 - molecule equilibrium polarizability
E0 - maximum electric field
ν0 & ν - excitation and vibrational frequency, respectively
rmax - maximum vibrational amplitude
’Modulation’ of linear optical polarizabiliy due to presence of internal
vibrations
Intensity
Values at
~ 1500 cm-1
I(l) >> IS >> IaS
1 >> 10-6 >> 10-9
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Mechanism of Enhancement in SERS
Chemical Mechanism
Electromagnetic
Mechanism
Electromagnetic Mechanism
Mechanism
Electromagnetic
Induced dipole moment in a molecule
p   Elocal
The molecular polarizability increases
due to the formation of charge transfer
(CT) metal-molecule complex. The CT
band is in resonance with both the
incident and scattered light.
Enhancement Contribution: 102-103
The local electric field experienced by
the adsorbed molecule increases due to
excitation of localized surface plasmons
by both the incident and scattered
photons
Enhancement Contribution: 106-108
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 SERS depends on the local field to the fourth power (Elocal ) in the vicinity of
an adsorbate molecule.
 It is a small range effect (~ 2-3 nm).
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Endocrine System
Endocrine system is an ensemble of glands which secrete different types of
hormones directly into the blood stream to maintain homeostasis. The endocrine
system regulates many vital processes in a living organism, such as development,
growth, reproduction and metabolism.
Endocrine Disruptors
Extrogeneous agents that interfere with the production, release, transport,
metabolism, binding, action or elimination of natural hormones in the body
responsible for the maintenance of homeostasis and regulation of developmental
processes
Possible Factors
Chemical /Industrial waste, Natural reasons/factors
Preservatives, processed food, unbalanced metabolism Pesticides, etc.
Detection of Vg is important to understand the estrogenic contamination of
aquatic environment.
 Representative of a kind of pollution in water/aquatic species
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