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Workshop Biosensor Brief


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characterization and functionalities of SAMs based biosensors

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Workshop Biosensor Brief

  1. 1. Characterisation of Biosensors Oxford Workshop 4-5 July 2009 Post Grad Cert Nanotechnology Nasrin Azadeh-McGuire University of Oxford Begbroke Science Park & Dept of Continuing Education
  2. 2. Bioaffinity Based Biosensor and immunosensor <ul><li>Biosensor is a detection device with high analytical specificity made of layers of biological elements with biorecognition quality that are immobilized on a substrate having reversible interactions with the analyte and a signal transducer. </li></ul><ul><li>Immunosensors are biosensors that have antibodies as biological element. </li></ul>
  3. 3. Surface characterisation of biosensors <ul><li>Chemical analysis to confirm covalent Immobilization </li></ul><ul><li>Surface density to detect distribution and location </li></ul><ul><li>Structural analysis to inform spatial relation and anatomy </li></ul><ul><li>Functional information about biorecognition </li></ul>
  4. 4. RMS roughness of Si wafer AFM <ul><ul><li>Roughness of a surface concerns topographical measure that occurs in the form of digs, particles, pits, granularity, deposited crystallites, and undulations on silicon wafers. </li></ul></ul><ul><ul><li>AFM tip’s radius of curvature at nanoscale makes it the technique of choice for roughness analysis </li></ul></ul><ul><ul><li>AFM in normal contact topographical mode </li></ul></ul><ul><ul><li>surface roughness measured by statistical account of peaks and valleys that gives the RMS ( root-mean-square) , an average of height deviations of sample points . </li></ul></ul><ul><ul><li>δ = Ö 1/N ∑ ( z i -μ) ² </li></ul></ul><ul><ul><li>N = number of data points, Z i = surface height variation at i th point , μ= average height </li></ul></ul>
  5. 5. XPS for determining Surface Silicon Oxide Layer & Contamination <ul><li>XPS measures elemental composition of top atomic layers (except H and He) </li></ul><ul><li>Elements unique binding energy are identifiable making XPS easy to read </li></ul><ul><li>XPS investigates Chain mobility, Contamination, Surface modification, and Chemical reactions, and Chemical states of the surfaces </li></ul><ul><li>Mono-energetic X-rays radiation cause emission of photoelectrons PE </li></ul><ul><li>Measuring PEs binding energy, intensity of their peak, elemental identity and quantity </li></ul>
  6. 6. SAMs <ul><li>Self assembled monolayers SAMs are long chain monolayers of diverse functionalities organised on surface of noble metals or silane for use as sensor arrays </li></ul><ul><li>Two main coupling schemes </li></ul><ul><li>thiol coupling </li></ul><ul><li>silane coupling </li></ul>
  7. 7. Silane coupling <ul><li>Silane coupling is covalent coupling onto a cross-linked surface shell </li></ul><ul><li> Organosilanes bind to the surface hydroxyl groups through Si-O bonds </li></ul>
  8. 8. Thiolated Au film coupling <ul><li>First monolayer appears within few minutes after immersion in dilute solution of alkanothiols in ethanol </li></ul><ul><li>After ~ 12 hours the film is highly packed by van der Waals forces , contaminations are replaced, defects are filled </li></ul><ul><li>Chemisorption of thiols onto gold surfaces mostly has covalent character </li></ul><ul><li>Au oxide surface layer reduced by the thiol form covalent link to Thiol monolayers </li></ul>
  9. 9. Surface Characterisation of SAMs <ul><li>ToF SIMS with elemental and molecular fingerprinting; detect distribution and location of biomolecules </li></ul><ul><li>Tip enhanced Raman Spectroscopy , FT-Raman Spectroscopy for molecular recognition </li></ul><ul><li>AFM tapping mode for imaging biofunctionalised surfaces, high resolution, ability to operate in liquid medium, topographical information, images of individual molecule measure molecular dimensions </li></ul><ul><li>AFM as force sensing tool ; AFM with functionalised tip, f-d mode probes functionality, elastic property, adhesion strength, and range of inter-atomic/molecular interactions for molecular recognition </li></ul><ul><li>XPS complemented with Ellipsometric and contact angle measurement can confirm the presence of immobilized molecules </li></ul>
  10. 10. ToF SIMS <ul><li>SIMS analyses mass of ions and molecular fragments ejected from surface (dept of 10-20 Å ) </li></ul><ul><li>TOF-SIMS: pulse primary ion beam at low fluence to ionize and desorbe the sample surface </li></ul><ul><li>ToF analyzer detects time of flight of secondary ions to mass spectrometer with their different velocities </li></ul><ul><li>Mass spectrum counts of secondary ions emitted to determine molecular species. </li></ul><ul><li>Highly focused primary ion beam (ca 1 μ dia ) images visualising molecular distribution on the surface </li></ul><ul><li>TOF-SIMS spectrum of (PET) (mass peaks for the fragments of a PET molecule) </li></ul>
  11. 11. ToF SIMS <ul><li>Advantageous: </li></ul><ul><li>Mass spectra with high mass resolution and high mass range (several thousand Daltons) </li></ul><ul><li>Molecular characterisation of surfaces, functionalities, and oxidation states </li></ul><ul><li>Limitations: </li></ul><ul><li>requires substantial interpretations of information rich spectra </li></ul><ul><li>Peak overlaps and interference </li></ul><ul><li>Larger molecules produce many smaller fragment ions </li></ul>
  12. 12. Microcantilevers as biological sensors with microscale length and width and nanoscale thickness <ul><li>Microlever Dynamic (resonance) mode measures resonant frequency changes due to added mass </li></ul><ul><li>Microlever Static (deflection) mode measures deflection of lever, displacement of the tip due to changes of stress or mass loading (operates both in air and liquid) </li></ul>
  13. 13. Functionalised micro-lever sensors <ul><li>Measuring deflection of lever (Static Mode) </li></ul><ul><li>piezoresistive </li></ul><ul><li>capacitive </li></ul><ul><li>optical reflection </li></ul><ul><li>piezoelectric </li></ul><ul><li>Measuring resonant frequency (Dynamic mode) </li></ul><ul><li>piezoelectric - oscillations with measurable amplitude </li></ul><ul><li>magnetically actuated photothermal – using diode laser </li></ul><ul><li>acoustic technique – using sound waves </li></ul><ul><li> bending responses due to various concentration of streptavidin </li></ul>
  14. 14. References <ul><li>Bruno Pettinger, Tip-enhanced Raman scattering: Influence of the tip-surface geometry on optical resonance and enhancement, Surface Science 603 (2009) 1335–1341 </li></ul><ul><li>Myhra S, A review of enabling technologies based on scanning probe microscopy relevant to bioanalysis, Biosensors and Bioelectronics, Volume 19, Issue 11, 15 June 2004, Pages 1345-1354 </li></ul><ul><li>Matthias Rief, Single Molecule Force Spectroscopy on Polysaccharides by Atomic Force Microscopy, Science, Vol. 275. no. 5304, pp. 1295 - 129 </li></ul><ul><li>Emanuele Ostuni, (Harvard Univ), The interaction of proteins and cells with self-assembled monolayers of alkanethiolates on gold and silver, Colloids and Surfaces B: Biointerfaces, Volume 15, Issue 1, 31 August 1999, Pages 3-30 </li></ul><ul><li>Wagner P, (Stanford Uni), Immobilization strategies for biological scanning probe microscopy, Volume 430, Issues 1-2, 23 June 1998, Pages 112-115 </li></ul><ul><li>Wang C et al, Ultrasensitive biochemical sensors based on microcantilevers of atomic force microscope, Analytical Biochemistry, Volume 363, Issue 1, 1 April 2007, Pages 1-11 </li></ul><ul><li>N Sandhyarani and T Pradeep, Characteristics of alkanethiol self assembled monolayers prepared on sputtered gold films: a surface enhanced Raman spectroscopic investigation, Vacuum, Volume 49, Issue 4, April 1998, Pages 279-284 </li></ul><ul><li>Ijeoma M. Nnebe, Carnegie Univ, Dynamic Atomic Force Microscopy Studies to Characterize Heterogeneous Surfaces, </li></ul><ul><li>Henke L et al, An AFM determination of the effects on surface roughness caused by cleaning of fused silica and glass substrates in the process of optical biosensor preparation, Biosensors and Bioelectronics 17 (2002) 547_/555 </li></ul><ul><li>Urban G, Micro- and nanobiosensors—state of the art and trends, Measurement Science and Technology 20 (2009) 012001 (18pp) </li></ul><ul><li>Louis Tiefenauer, Biointerface analysis on a molecular level: New tools for biosensor research, Colloids and Surfaces B: Biointerfaces,, Volume 23, Issues 2-3, February 2002, Pages 95-114 </li></ul><ul><li>Yam Chi Ming et al, Preparation, characterization, resistance to protein adsorption, and specific avidin–biotin binding of poly(amidoamine) dendrimers functionalized with oligo(ethylene glycol) on gold, Journal of Colloid and Interface Science, Volume 296, Issue 1, April 2006, Pages 118-130 </li></ul><ul><li> </li></ul><ul><li> </li></ul><ul><li> </li></ul><ul><li> </li></ul>