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    Surface Plasmon Resonance: Antigen-Antibody Interactions: Vamsi K. Mudhivarthi

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    Surface plasmon resonance (SPR) is a technique that can rapidly monitor biomolecular interactions in real-time. It works by detecting changes in the refractive index near a gold surface when molecules bind to it. This allows it to study interactions like antigen-antibody binding through both association and dissociation phases. SPR has applications in physical adsorption processes as well as biological areas like protein structure determination, epitope mapping, and tissue engineering. While sensitive, it has limitations with low molecular weight compounds and rate effects from mass transport.

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    Surface Plasmon Resonance: Antigen-Antibody Interactions: Vamsi K. Mudhivarthi

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    Gobi
    44 views21 pages
    Surface plasmon resonance (SPR) is a technique that can rapidly monitor biomolecular interactions in real-time. It works by detecting changes in the refractive index near a gold surface when molecules bind to it. This allows it to study interactions like antigen-antibody binding through both association and dissociation phases. SPR has applications in physical adsorption processes as well as biological areas like protein structure determination, epitope mapping, and tissue engineering. While sensitive, it has limitations with low molecular weight compounds and rate effects from mass transport.

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    Biomaterials characterization

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    Vamsi

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    Surface plasmon resonance (SPR) is a technique that can rapidly monitor biomolecular interactions in real-time. It works by detecting changes in the refractive index near a gold surface when molecules bind to it. This allows it to study interactions like antigen-antibody binding through both association and dissociation phases. SPR has applications in physical adsorption processes as well as biological areas like protein structure determination, epitope mapping, and tissue engineering. While sensitive, it has limitations with low molecular weight compounds and rate effects from mass transport.

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    44 views21 pages

    Surface Plasmon Resonance: Antigen-Antibody Interactions: Vamsi K. Mudhivarthi

    Uploaded by

    Gobi
    Surface plasmon resonance (SPR) is a technique that can rapidly monitor biomolecular interactions in real-time. It works by detecting changes in the refractive index near a gold surface when molecules bind to it. This allows it to study interactions like antigen-antibody binding through both association and dissociation phases. SPR has applications in physical adsorption processes as well as biological areas like protein structure determination, epitope mapping, and tissue engineering. While sensitive, it has limitations with low molecular weight compounds and rate effects from mass transport.

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    Surface Plasmon Resonance:

    antigen-antibody interactions

    Vamsi K. Mudhivarthi
    Analytical Techniques for Biomaterial
    Characterization
    • Scanning Probe Microscopy (SPM):
    – image changes at surface

    • Attenuated Total Reflectance Infrared Spectroscopy (ATR-IR):


    – study conformational changes at solid-aqueous interfaces,
    although lacks sensitivity

    • Spectral Ellipsometry:
    – determination of thickness and refractive index of adsorbed
    layer

    • Surface Plasmon Resonance (SPR):


    – rapidly monitor dynamic processes to a wide range of
    biomedically relevant interfaces.
    What is SPR?

    • Surface Plasmon: Longitudinal charge density wave along the


    interface of two media, where one is metal and other is dielectric
    SPR Principle

    • Linear relationship is found between resonance energy and


    mass concentration of biochemically relevant molecules.
    Advantage of using gold film

    • Gold: Non-magnetic, surface plasmon wave is p-polarized, and


    due to its electromagnetic and surface propagating nature, creates
    enhanced evanescent wave
    Gold-thiol Chemistry
    Typical SPR Signal
    SPR: Kinetics of Association phase
    Ka
    A s urface + B AB
    Kd

    Km

    A bulk

    dR/dt = KaC (Rmax - R) - KdR


    or
    dR/dt = KaC Rmax - (Kd+ KaC)R
    • C= Concentration of analyte
    • Rmax = maximum analyte binding capacity of the surface in RU
    • R = SPR signal at time t in RU
    SPR: Kinetics of Association phase

    • Kd is not very reliable as KaC >>Kd


    SPR: Kinetics of Dissociation phase

    dR/dt = - KdR

    After integration and logarithm


    we have
    ln (R0/Rt) = Kd (t-t 0)

    • Rt is response at time t in RU
    • R0 is response at an arbitrary starting point
    Applications of SPR
    • Physical applications: measure dielectric properties, adsorption
    processes, surface degradation or hydration of
    – Thin organic monolayers or bilayers
    – Polymer films

    • Biological applications: as biosensors for specific biological


    interactions including adsorption and desorption kinetics, antigen-
    antibody binding and epitope mapping for determination of
    – Biomolecular structure and interactions of proteins, DNA &
    Viruses
    – Lipid Bilayers
    – Non-specific biomolecular interactions-bio-compatibility
    – Tissue engineering
    SPR: Physical applications

    Thin organic monolayers or bilayers Polymer films


    SPR: Biological applications

    Epitope mapping Tissue engineering


    Immune response to antigen
    Antibody structure
    Single step analysis: FMDV antigen-antibody
    interactions

    Ka = 9.0 * 104 M-1 S-1


    Kd = 1.2 * 10-3 S-1
    Analysis of the antigen of different strains
    binding to antibody: single step
    Analysis of the antigen of different strains
    binding to antibody: single step
    Advantage of SPR

    • Ability to perform real-time measurement:


    – Insight to dynamic nature of binding system and layer
    formation

    • Use of selective slides to study binding events:


    – Eliminate the need for labeled reactants

    • Exceptional sensitivity:
    – Small quantities of purified reagents are required
    Disadavantages

    • Disadvantage of SPR:
    – Lack of sensitivity when monitoring low molecular
    weight adsorbates
    – Rate limiting factor of mass transport-affecting kinetic
    analysis

    • Methods to improve sensitivity:


    – Coupling to AFM
    – Coupling with Mass-spectrometry
    References

    • Green R. J., Frazier R. A., et. al., Biomaterials 21 (2000) 1823-1835.


    • http://brahms.chem.uic.edu/~cgpage/research/sprintro.html
    • Biacore Manual
    • David Andreu et. al., Journal of immunological methods 235 (2000)
    101-111.
    • A. McGill et. al., Journal of immunological methods 297 (2005) 143-
    152.
    • Marc H. V. Van Regnmortel et. al., Journal of molecular recognition
    11 (1998) 163-167.
    • Daniele Altschuh et. al., Biochemistry 31 (1992) 6298-6304.

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