Surface plasmon resonance imaging is used to detect binding events between biological molecules on gold surfaces without labeling. SPR imaging works by exciting surface plasmons at a gold-liquid interface using p-polarized light. Changes in the local refractive index from molecule binding cause shifts in the resonance angle that can be detected using a CCD camera. In the presented experiment, SPR imaging was used to obtain real-time images showing the immobilization of thiol-modified DNA probes on a gold surface and their subsequent hybridization with complementary DNA sequences in liquid. SPR imaging provides label-free detection with high sensitivity and the ability to monitor binding kinetics in situ.
Surface Plasmon Resonance (SPR) and its ApplicationDr. Barkha Gupta
DR. BARKHA GUPTA
ASSISTANT PROFESSOR (VETERINARY BIOCHEMISTRY)
DEPARTMENT OF VETERINARY PHYSIOLOGY AND BIOCHEMISTRY
POST GRADUATE INSTITUTE OF VETERINARY EDUCATION AND RESEARCH (PGIVER), JAIPUR RAJASTHAN UNIVERSITY OF VETERINARY AND ANIMAL SCIENCES (RAJUVAS), BIKANER
YouTube Channel: Barkha’s Vet Sphere
Surface Plasmon Resonance (SPR) and its ApplicationDr. Barkha Gupta
DR. BARKHA GUPTA
ASSISTANT PROFESSOR (VETERINARY BIOCHEMISTRY)
DEPARTMENT OF VETERINARY PHYSIOLOGY AND BIOCHEMISTRY
POST GRADUATE INSTITUTE OF VETERINARY EDUCATION AND RESEARCH (PGIVER), JAIPUR RAJASTHAN UNIVERSITY OF VETERINARY AND ANIMAL SCIENCES (RAJUVAS), BIKANER
YouTube Channel: Barkha’s Vet Sphere
Introduction to nanoscience and nanotechnologyaimanmukhtar1
Introduction of nanoscience/nanotechnology ,properties/potential applications of nanomaterials and electrodeposition of metal single component and alloy nanowires in AAO template
X ray crystallography to visualize protein structure.Ritam38
This ppt discusses in detail the process of X ray Crystallography.
Made by the following 3rd year Bs-Ms students of IISER Kolkata:
B Sri Sindhu
Rasiwala Hassan Shabbir
Ritam Samanta
Himanshu Gupta
Sakshi Ajay Shrisath
Aditya Borkar
Diana Denzil Fernandez
Neha Kumari
.Sowmya
Anjali Mohan
Debanjana Mondal
Aanandita Gope
Shruti Santosh Sail
Carbohydrate Research 405 (2015) 55–65Contents lists availab.docxwendolynhalbert
Carbohydrate Research 405 (2015) 55–65
Contents lists available at ScienceDirect
Carbohydrate Research
j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / c a r r e s
Electrochemical synthesis of nanostructured gold film for the study
of carbohydrate–lectin interactions using localized surface plasmon
resonance spectroscopy
http://dx.doi.org/10.1016/j.carres.2014.08.019
0008-6215/� 2014 Elsevier Ltd. All rights reserved.
⇑ Corresponding author. Tel.: +1 (314) 516 5346; fax: +1 (314) 516 5342.
E-mail address: [email protected] (K.J. Stine).
Jay K. Bhattarai a,b, Abeera Sharma a,b, Kohki Fujikawa a, Alexei V. Demchenko a, Keith J. Stine a,b,⇑
a Department of Chemistry and Biochemistry, University of Missouri—St. Louis, One University Boulevard, St. Louis, MO 63121, United States
b Center for Nanoscience, University of Missouri—St. Louis, One University Boulevard, St. Louis, MO 63121, United States
a r t i c l e i n f o a b s t r a c t
Article history:
Received 30 April 2014
Received in revised form 15 July 2014
Accepted 30 August 2014
Available online 16 September 2014
Keywords:
Lectin
Mannose
Monolayer
Plasmon
Gold
Localized surface plasmon resonance (LSPR) spectroscopy is a label-free chemical and biological molec-
ular sensing technique whose sensitivity depends upon development of nanostructured transducers.
Herein, we report an electrodeposition method for fabricating nanostructured gold films (NGFs) that
can be used as transducers in LSPR spectroscopy. The NGF was prepared by electrodepositing gold from
potassium dicyanoaurate solution onto a flat gold surface using two sequential controlled potential steps.
Imaging by scanning electron microscopy reveals a morphology consisting of randomly configured block-
like nanostructures. The bulk refractive index sensitivity of the prepared NGF is 100 ± 2 nm RIU�1 and the
initial peak in the reflectance spectrum is at 518 ± 1 nm under N2(g). The figure of merit is 1.7. In addition,
we have studied the interaction between carbohydrate (mannose) and lectin (Concanavalin A) on the
NGF surface using LSPR spectroscopy by measuring the interaction of 8-mercaptooctyl-a-D-mannopyran-
oside (aMan-C8-SH) with Concanavalin A by first immobilizing aMan-C8-SH in mixed SAMs with 3,6-
dioxa-8-mercaptooctanol (TEG-SH) on the NGF surface. The interaction of Con A with the mixed SAMs
is confirmed using electrochemical impedance spectroscopy. Finally, the NGF surface was regenerated
to its original sensitivity by removing the SAM and the bound biomolecules. The results from these exper-
iments contribute toward the development of inexpensive LSPR based sensors that could be useful for
studying glycan–protein interactions and other bioanalytical purposes.
� 2014 Elsevier Ltd. All rights reserved.
1. Introduction
Localized surface plasmon resonance (LSPR) spectroscopy based
on the development of noble metal nanostructures with tunable
and responsive plasmonic behavior has become of b ...
NMR, principle and instrumentation by kk sahu sirKAUSHAL SAHU
Introduction
History
Principle
Assembly
Solvents
Chemical shift
Factors affecting chemical shift
2D NMR
NOE effect
NOESY
COSY
Application
Conclusion
References
Introduction to nanoscience and nanotechnologyaimanmukhtar1
Introduction of nanoscience/nanotechnology ,properties/potential applications of nanomaterials and electrodeposition of metal single component and alloy nanowires in AAO template
X ray crystallography to visualize protein structure.Ritam38
This ppt discusses in detail the process of X ray Crystallography.
Made by the following 3rd year Bs-Ms students of IISER Kolkata:
B Sri Sindhu
Rasiwala Hassan Shabbir
Ritam Samanta
Himanshu Gupta
Sakshi Ajay Shrisath
Aditya Borkar
Diana Denzil Fernandez
Neha Kumari
.Sowmya
Anjali Mohan
Debanjana Mondal
Aanandita Gope
Shruti Santosh Sail
Carbohydrate Research 405 (2015) 55–65Contents lists availab.docxwendolynhalbert
Carbohydrate Research 405 (2015) 55–65
Contents lists available at ScienceDirect
Carbohydrate Research
j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / c a r r e s
Electrochemical synthesis of nanostructured gold film for the study
of carbohydrate–lectin interactions using localized surface plasmon
resonance spectroscopy
http://dx.doi.org/10.1016/j.carres.2014.08.019
0008-6215/� 2014 Elsevier Ltd. All rights reserved.
⇑ Corresponding author. Tel.: +1 (314) 516 5346; fax: +1 (314) 516 5342.
E-mail address: [email protected] (K.J. Stine).
Jay K. Bhattarai a,b, Abeera Sharma a,b, Kohki Fujikawa a, Alexei V. Demchenko a, Keith J. Stine a,b,⇑
a Department of Chemistry and Biochemistry, University of Missouri—St. Louis, One University Boulevard, St. Louis, MO 63121, United States
b Center for Nanoscience, University of Missouri—St. Louis, One University Boulevard, St. Louis, MO 63121, United States
a r t i c l e i n f o a b s t r a c t
Article history:
Received 30 April 2014
Received in revised form 15 July 2014
Accepted 30 August 2014
Available online 16 September 2014
Keywords:
Lectin
Mannose
Monolayer
Plasmon
Gold
Localized surface plasmon resonance (LSPR) spectroscopy is a label-free chemical and biological molec-
ular sensing technique whose sensitivity depends upon development of nanostructured transducers.
Herein, we report an electrodeposition method for fabricating nanostructured gold films (NGFs) that
can be used as transducers in LSPR spectroscopy. The NGF was prepared by electrodepositing gold from
potassium dicyanoaurate solution onto a flat gold surface using two sequential controlled potential steps.
Imaging by scanning electron microscopy reveals a morphology consisting of randomly configured block-
like nanostructures. The bulk refractive index sensitivity of the prepared NGF is 100 ± 2 nm RIU�1 and the
initial peak in the reflectance spectrum is at 518 ± 1 nm under N2(g). The figure of merit is 1.7. In addition,
we have studied the interaction between carbohydrate (mannose) and lectin (Concanavalin A) on the
NGF surface using LSPR spectroscopy by measuring the interaction of 8-mercaptooctyl-a-D-mannopyran-
oside (aMan-C8-SH) with Concanavalin A by first immobilizing aMan-C8-SH in mixed SAMs with 3,6-
dioxa-8-mercaptooctanol (TEG-SH) on the NGF surface. The interaction of Con A with the mixed SAMs
is confirmed using electrochemical impedance spectroscopy. Finally, the NGF surface was regenerated
to its original sensitivity by removing the SAM and the bound biomolecules. The results from these exper-
iments contribute toward the development of inexpensive LSPR based sensors that could be useful for
studying glycan–protein interactions and other bioanalytical purposes.
� 2014 Elsevier Ltd. All rights reserved.
1. Introduction
Localized surface plasmon resonance (LSPR) spectroscopy based
on the development of noble metal nanostructures with tunable
and responsive plasmonic behavior has become of b ...
NMR, principle and instrumentation by kk sahu sirKAUSHAL SAHU
Introduction
History
Principle
Assembly
Solvents
Chemical shift
Factors affecting chemical shift
2D NMR
NOE effect
NOESY
COSY
Application
Conclusion
References
(May 29th, 2024) Advancements in Intravital Microscopy- Insights for Preclini...Scintica Instrumentation
Intravital microscopy (IVM) is a powerful tool utilized to study cellular behavior over time and space in vivo. Much of our understanding of cell biology has been accomplished using various in vitro and ex vivo methods; however, these studies do not necessarily reflect the natural dynamics of biological processes. Unlike traditional cell culture or fixed tissue imaging, IVM allows for the ultra-fast high-resolution imaging of cellular processes over time and space and were studied in its natural environment. Real-time visualization of biological processes in the context of an intact organism helps maintain physiological relevance and provide insights into the progression of disease, response to treatments or developmental processes.
In this webinar we give an overview of advanced applications of the IVM system in preclinical research. IVIM technology is a provider of all-in-one intravital microscopy systems and solutions optimized for in vivo imaging of live animal models at sub-micron resolution. The system’s unique features and user-friendly software enables researchers to probe fast dynamic biological processes such as immune cell tracking, cell-cell interaction as well as vascularization and tumor metastasis with exceptional detail. This webinar will also give an overview of IVM being utilized in drug development, offering a view into the intricate interaction between drugs/nanoparticles and tissues in vivo and allows for the evaluation of therapeutic intervention in a variety of tissues and organs. This interdisciplinary collaboration continues to drive the advancements of novel therapeutic strategies.
Professional air quality monitoring systems provide immediate, on-site data for analysis, compliance, and decision-making.
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The increased availability of biomedical data, particularly in the public domain, offers the opportunity to better understand human health and to develop effective therapeutics for a wide range of unmet medical needs. However, data scientists remain stymied by the fact that data remain hard to find and to productively reuse because data and their metadata i) are wholly inaccessible, ii) are in non-standard or incompatible representations, iii) do not conform to community standards, and iv) have unclear or highly restricted terms and conditions that preclude legitimate reuse. These limitations require a rethink on data can be made machine and AI-ready - the key motivation behind the FAIR Guiding Principles. Concurrently, while recent efforts have explored the use of deep learning to fuse disparate data into predictive models for a wide range of biomedical applications, these models often fail even when the correct answer is already known, and fail to explain individual predictions in terms that data scientists can appreciate. These limitations suggest that new methods to produce practical artificial intelligence are still needed.
In this talk, I will discuss our work in (1) building an integrative knowledge infrastructure to prepare FAIR and "AI-ready" data and services along with (2) neurosymbolic AI methods to improve the quality of predictions and to generate plausible explanations. Attention is given to standards, platforms, and methods to wrangle knowledge into simple, but effective semantic and latent representations, and to make these available into standards-compliant and discoverable interfaces that can be used in model building, validation, and explanation. Our work, and those of others in the field, creates a baseline for building trustworthy and easy to deploy AI models in biomedicine.
Bio
Dr. Michel Dumontier is the Distinguished Professor of Data Science at Maastricht University, founder and executive director of the Institute of Data Science, and co-founder of the FAIR (Findable, Accessible, Interoperable and Reusable) data principles. His research explores socio-technological approaches for responsible discovery science, which includes collaborative multi-modal knowledge graphs, privacy-preserving distributed data mining, and AI methods for drug discovery and personalized medicine. His work is supported through the Dutch National Research Agenda, the Netherlands Organisation for Scientific Research, Horizon Europe, the European Open Science Cloud, the US National Institutes of Health, and a Marie-Curie Innovative Training Network. He is the editor-in-chief for the journal Data Science and is internationally recognized for his contributions in bioinformatics, biomedical informatics, and semantic technologies including ontologies and linked data.
5. How does SPR work?
http://www.biochem.mpg.de/oesterhelt/xlab/spfs.html
• This Kretschmann
Experimental System
uses a metal film thin
enough to monitor the
plasmon
A plasmon can be thought of as a
ray of light bound onto a surface -
propagating among the surface
and presenting itself as an
electromagnetic field.
6. Otto Experimental System (1968)
•Impossible to
observe plasmon
through infinite metal
•The Kretschmann
system (1971) is
used in designs of
SPR instruments
7. Metals
• Gold and silver are most commonly used.
Aluminum and copper may also be
suitable metals.
8. SPR
• A surface-plasmon-resonance is excited at a
metal-dielectric interface by a monochromatic, p-
polarized light beam, such as He-Ne laser beam
• The surface plasmon is sensitive to changes in
the environment near the interface and therefore
has potential as a sensing probe.
• Sensitive detection method that monitors
variations in thickness and refractive index in
ultra-thin films
9. More about SPR
• It is a non-destructive means of sensing
• Surface Plasmons have already been
used for gas sensing, biosensing,
immuno-sensing and electrochemical
studies.
10. Modifications for
SPR imaging apparatus
• The p-polarized He-Ne laser beam passes
through a spatial filter and can be
expanded using a beam expander
• The reflected light is collected by a CCD
camera to produce an “SPR image”
11. SPR imaging apparatus
• Spatially-filtered,
expanded, p-polarized
HeNe laser beam
illuminates the gold sample
through a prism coupler.
• Reflected light from the
gold surface, containing the
SPR image, is monitored
with a CCD camera.
• The angle of incidence can
be changed by rotating the
entire sample assembly.
A.J. Thiel et. al., Anal. Chem. 69 (1997), pp. 4948–4956.
12. SPR Imaging Experiment
• Investigation of adsorption processes of
oligonucleotides onto gold substrates in aqueous
buffer solution
• Monitored hybridization process of thiol-modified
single stranded oligonucleotide anchored to gold
surface with its complementary sequence
• Traditional analysis of hybridization involves
labeling, such as radioactive isotopes or
fluorescent molecules
13. Preparation
• Glass slides (cleaned in piranha solution)
• 2nm chromium layer (adhesion layer) followed
by 50nm gold film
• Gold surface spotted with HS-ssDNA (thiolated
oligonucleotide probes) and left to react for at
least 18h
• After soaked and rinsed in water, the probes
were ready for hybridization
• Probes were immersed in hybridization buffer
containing complementary oligonucleotide in
Phosphate buffer system at pH 7.4
14. Hybridized helices formed on gold substrate
•R. Rella, et al. Biosensors and Bioelectronics. 20 (2004), pp.1140-1148.
•Thiol-modified, single
stranded oligonucleotide
anchored to gold surface
•Immersed in
mercaptoethanol
for 18h to eliminate
aspecific
adsorption sites on
the gold surface
•Forms dsDNA with
complementary sequence
15. SPR Reflectivity
•The cusp does not change
in the presence of the
oligo, therefore ensures
reproducibility of the
scanning measurement
•0.41 degree shift of the
self assembled monolayer
from the bare gold surface
was used to estimate a
5nm thickness of self
assembled oligonucleotide
and refractive index of 1.65
•R. Rella, et al. Biosensors and Bioelectronics. 20 (2004), pp.1140-1148.
16. 2D and 3D Images
of ssDNA
•Shows the 5 spots of self
assembled thio-
oligonucleotide DNA probes
immobilized on the gold
surface
•Color variation indicates
variation in the thickness of
the self assembled
monolayer (SAM)
•R. Rella, et al. Biosensors and Bioelectronics. 20 (2004), pp.1140-1148.
17. 3D Images of
ssDNA (top) and
dsDNA (bottom)
•SPR images of
immobilization (ssDNA)
and hybridization
(dsDNA) in liquid.
•Hybridization process
shown by variation in
resonance signal of a
specific self assembled
spot.
•R. Rella, et al. Biosensors and Bioelectronics. 20 (2004), pp.1140-1148.
18. 2D Images of
ssDNA (top) and
dsDNA (bottom)
•The increase in resonance
signal corresponds to the
increase in layer thickness
indicative of the hybridization
process
•R. Rella, et al. Biosensors and Bioelectronics. 20 (2004), pp.1140-1148.
19. Summary
• SPR imaging used to obtain images of
thio-oligonucleotides linked onto a gold
surface
• Detection of DNA hybridization using liquid
phase SPR imaging techniques
20. Advantages of SPR imaging
• Identification of binding events with label-free molecules
• High speed and sensitivity with real-time reaction
monitoring
• High spatial resolutions
• Identity specific vs. non-specific adsorption processes
• In-situ capabilities
21. Future Applications
• Using the technique to obtain the total number of
target DNA hybridization events per unit area
• Construct patterned gold substrates and self
assemble DNA probes of different nature
• Construct a DNA array suitable to detect specific
DNA sequence target
25. SIGNAL DETECTION
• The light source for SPR is a high efficiency near-
infrared light emitting diode which has a fixed range of
incident angles. The SPR response is monitored by a
fixed array of light sensitive diodes covering the whole
wedge of reflected light. The angle at which minimum
reflection occurs is detected and converted to the
resonance units. The SPR angle depends on several
factors, most notably the refractive index into which the
evanescent wave propagates on the non-illuminated side
of the surface. In addition the other parameters are the
wavelength of the incident light and the properties of the
metal film.
26. •As we change the angle of incidence of light (at a fixed frequency), at
some angle the projection of the light wave vector onto the metal film
becomes equal to the wave vector of SP's at the same frequency. Light
energy can then be effectively transferred to the SP's.
•If we vary the thickness of the metal film, the phase difference
between the reflected light and the light re-radiated by SP's will change.
If we scan the angle of incidence of light onto a metal film of the right
thickness, then at a certain angle the reflected light intensity will go
sharply to almost zero, indicating resonant coupling to surface
plasmons. This angle is always greater than the angle of total internal
reflection off the prism/outer dielectric interface, and is called the
attenuated total reflection (ATR) angle.
•And this angle is important, because the position and the width of this
‘resonance point’ are very sensitive to the properties of the surface and
the media next to it. It makes it possible to use surface plasmon
resonance techniques for chemical and biological sensing.
How it works
27. SPR imaging
• SPR imaging is a surface sensitive optical technique for monitoring the adsorption of
solution species into patterned molecular microarrays that have been made on
chemically modified metal surfaces (gold). SPR can be used to study the specific
adsorption of biopolymers (oligonucleotides, proteins, antibodies) onto DNA or protein
microarrays. The microarray format makes the SPR technique a high throughput
differential binding measurement system that can be used to accurately measure the
specific organic and biomolecular interactions with a variety of surfaces under
identical solution adsorption conditions.
• Various biomolecular interactions can be characterized including DNA-DNA, RNA-
DNA, protein-DNA and protein-protein binding. As a surface-sensitive measurement,
SPR is an indispensable tool for the study of in-situ biological systems of weakly
bound interactions. Since the SPR imaging experiment senses the change in the
local index of refraction upon adsorption onto the metal surface, no labeling of the
biomolecules is required. The combined simplicity and versatility of surface plasmon
resonance imaging measurements make them ideally suited to the study of ultra thin
organic films. The technique is extremely sensitive, provides reasonable lateral
resolution and can be used in both ex-situ and in-situ configurations to monitor the
real-time binding of label free analyte molecules to metal surfaces. The two
dimensional analysis of imaging data, combined with our microarray manufacturing
and processing technologies on gold surfaces makes surface plasmon resonance a
powerful tool for the parallel processing of multiplexed biomolecular interactions.
28. Specifications of the SPR Imager
• Sensitive to less than 1 angstrom molecular thickness range
• Liquid cell attachment for in-situ measurements
• High precision coupled prism and detector rotating stage
•
• Spectral range from 600nm to 1100nm
• Incident angle range for 40 to 70 degrees for differing prism and sampling systems
• High sensitivity near IR CCD detector with video interface
• Image acquisition interface and image processing software
• 110 or 220 volt versions