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Real Time Electrochemical Displacement Immunosensor for TCA detection

Real Time Electrochemical Displacement Immunosensor for TCA detection. María Viviana Duarte 2nd year PhD Student Project thesis course January 2004. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte. 5 W and 1 H.

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Real Time Electrochemical Displacement Immunosensor for TCA detection

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  1. Real Time Electrochemical Displacement Immunosensor for TCA detection María Viviana Duarte 2nd year PhD Student Project thesis course January 2004

  2. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte 5 W and 1 H Real Time Electrochemical Displacement Immunosensor for TCA detection - 2,4,6 trichloroanisole (TCA) detection using antiTCA-IgG What? Who? MV Duarte- 2nd year PhD student. Project thesis When? PhD studies Where? BBG group – Etseq Universitat Rovira I Virgili Why? TCA - as an indicator of contaminants responsible for cork taint How? Our Tools- Antibodies and Electrochemical immunosensor

  3. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte Why- An introduction Cost money to cork and wine industry Cork taint incidents An indicator! 2,4,6-trichloroanisole (TCA)

  4. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte How- Our Tools Immunosensors Selective immunodetection of TCA Antibodies Displacement methodology: reagentless and fast responding devices

  5. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte How- The detection Antibody antiTCA- IgG 150 k DA TCA 0,2 k DA

  6. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte

  7. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte t/seg 1- TCA- HRP Adsorption Amperometric signal e - substrate TCA-HRP substrate product Graphite electrode product HRP labelled TCA Materials and Methods- System 1

  8. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte Substrate addition t/seg TCA (analyte) additions e- substrate TCA-HRP substrate product Graphite electrode product HRP labelled TCA 2- TCA (analyte) addition Displacement 1- TCA- HRP Adsorption

  9. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte TCP or HaptenA TCA injections started System 1- Results Specific and non-specific Adsorption Adsorption Amperograms TCA- HRP on IgG electrode Displacement chronoamperogram Substrate • Substrate • TCA-HRP 1- TCA- HRP Adsorption 2- TCA addition

  10. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte t/seg 1- antimouse- HRP Adsorption Amperometric signal e - substrate Antimouse-HRP product substrate TCA-BSA product Materials and Methods- System 2

  11. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte e - substrate product substrate product 2- TCA (analyte) addition Displacement 1- Antimouse- HRP Adsorption

  12. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte BSA electrode System 2- Results BSA or TCA-BSA 5%- Co Pos 90%-PEG 5%; 30 min 37ºC anti TCA IgG; chornoamperometric signal of antimouse HRP Displacement experiments are the next step! TCA-BSA electrode

  13. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte 50 nm Incident light Reflected light 50 nm channel

  14. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte K Association=kon/koff K A (M-1) 7,31 E+5 1,90 E+5 kon: 5,22 E+2 M-1sec-1 + koff: 7,14 E-4 sec-1 Immobilized antiTCA antibody Hapten A- HRP kon: 8,97 E+2 M-1sec-1 + koff:4,72 E-3 sec-1 Immobilized antiTCA antibody Hapten 1- HRP

  15. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte Conclusions SPR Characterization: The affinity of immobilized AntiTCA IgG Clon 007 with Hapten A- HRP is 3,5 times higher than with Hapten1-HRP With the results obtained at the moment: • Immuno-electrochemical sensor: It seems that BSA- TCA electrodes (system 2) - AntiTCA IgG–antimouse HRP adsorbed could be use to study further TCA displacements

  16. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte Future work Antibody characterization- SPR Continue with the characterization of different anti TCA-IgG (clon) and their interaction with TCA – HRP conjugations, BSA- TCA conjugations, TCA, etc Immunoelectrochemical sensor Continue with electrochemichal System 2 and the Displacement experiments Use SPR information to improve System 1

  17. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte Thank you for your attention

  18. Real Time Electrochemical Displacement Immunosensor for TCA detection- MV Duarte Haptens TCA Hapten A Hapten 1

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