Electrochemical Scanning Tunneling Microscopy

Transcription

1 Electrochemical Scanning Tunneling Microscopy 1. An electrochemical cell usually contains three electrodes. The working electrode is the electrode whose properties are being studied. Current is passed between the working electrode and another electrode immersed in the solution, called the auxiliary or counter electrode. 2. The reference electrode in an electrochemical cell is usually one whose potential remains constant with time and can be prepared reproducibly. Common reference electrodes are the saturated calomel electrode (SCE) prepared from mercury in contact with mercurous chloride and saturated KCl solution, or a silver/silver chloride electrode in contact with saturated KCl. 3. The electrochemical characteristics of a working electrode are frequently obtained by observing how the faradic current varies with the potential of the electrode vs. the reference electrode, or voltammogram. 4. EC-STM cells are actually four-electrode cells, where both the tip and the substrate are working electrodes.

2 1. EC-STM can be used to characterize the solid-electrolyte interface at high spatial resolution. 2. The total current measured with the STM in an electrochemical environment is given by the sum of the tunneling current and the Faradic current, the latter results from electrochemical processes both at the substrate-electrolyte and the exposed tip-electrolyte interface. To reduce the unwanted contribution from the Faradic current to a small percentage, the exposed tip-electrolyte interfacial area has to be reduced. This can be achieved by coating the tip with an electrically insulating material, leaving only μm of the tip end exposed. The residual Faradic current can usually be kept below 100 pa. 3. Coating materials: glass, epoxy resin, Apiezon wax, polymer, or nail polish. 4. For EC-STM, it is essential to control the electrochemical reactions at the substrate and the exposed part of the tip by introducing a potentiostatic STM concept. This concept involves provision for independent adjustment of the potentials E S and E T of substrate and tip relative to a reference electrode (RE).

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4 1. Ideally, the cell should be closed to prevent evaporation of solution, which causes changes in the solution composition and promotes thermal drifts. However, this is incompatible with the use of an STM scanning head. 2. Most electrochemical experiments are carried out under an inert atmosphere by bubbling nitrogen or argon through the solution before an experiment and flowing these inert gases over the solution during the studies. This prevents oxygen and impurities in the atmosphere from entering the cell solution. 3. Most EC-STM experiments reported have used solutions exposed to the atmosphere. 4. Bipotentiostats are usually based on operational amplifier circuits and allow independent adjustment of E t and E s and measurement of both the tip current (I t ) and the substrate current (I s ). 5. EC-STM has been used as an effective tool for real-time, in-situ characterization of both single-crystalline and mesoscopic structures at the solid/electrolyte interface. Defects and their dynamics have been studied on an atomic scale basis. Nucleation and growth processes associated with surface reconstruction, adsorption/desorption, deposition/dissolution, passivation and other phenomena have been examined.

5 The Electric Double Layer Normalized ion densities for Na + (solid line) and Cl - ( ) near a positively (a) and a negatively (b) charged wall (z=0) in 2.2M NaCl. Angew. Chem. Int. Ed. 40, 1162 (2001).

6 Surface Reconstruction Au(100) surface in 0.1M H 2 SO 4 Unreconstructed Au(100) surface recorded shortly after lifting of the (hex) reconstruction +0.35V Potential-induced reconstruction

7 The Study of Adsorbates Angew. Chem. Int. Ed. 40, 1162 (2001). Pd on Au (underpotential deposition)

8 Self-Assembled Monolayer Reversible orderdisorder transition Ethanethiol on Au(111) in 0.1M H 2 SO 4 Angew. Chem. Int. Ed. 40, 1162 (2001).

9 500 nm x 500 nm Metal Deposition-Underpotential deposition of Pd Pd deposition onto Au(111) from 0.1 mm H 2 PdCl 4 in 0.1M H 2 SO 4

10 500 nm x 500 nm Surface Science 443, 19( nm x 550 nm 350 mv

11 Fabrication of Nanostructures with EC-STM Jump-to-contact between a Cu-covered STM tip and an Au substrate. Cu clusters on Au(111) 0.05M H 2 SO 4 +1mM CuSO 4 Science 248, 454 (1990) Generated by single voltage pulses of 2 ms duration to the z-piezo of an STM.

12 Nanoscale Electrodeposition J. Appl. Phys.87,7007(2000) Nernst equation: E = E 0 + (RT/nF) ln [M n+ ] 0.25 M Na 2 SO 4 and 1 mm CoSO 4 (ph~4 5, T=298K) Current voltage characteristics of the Co deposition and dissolution at the Au STM tip. The STM tip is in a constant distance of nm from the substrate surface; E WE =-770 mv is held constant during the complete deposition cycle.

13 Cantilever-Based Chemical and Biological Sensors silicon cantilever array Molecular recognition DNA hybridization Receptor-ligand binding Science 288, 316 (2000) surface stress changes

14 Chemical and Biological Sensors A.Static deformations B. Resonance operation COMMON READOUT SCHEMES 1. Optical methods 2. Piezoresistance method 3. Piezoelectric method 4. Capacitance method A. Gas phase analytes B. Liquid phase analytes Sensitivity & Selectivity (Specificity) REV.OF SCI. INSTR. 75, 2229 (2004)

15 Virus detection using nanoelectromechanical devices Mass sensitivities on the order of g/hz Detect the mass of single-virus particles l=6 μm, w=0.5 μ m, t=150 nm with a 1 μm x 1 μm paddle. Scale bar corresponds to 2 μm. Appl. Phys. Lett. 85, 2604 (2004)

16 The Scanning Probe Microscope in Biology 1. AFM experiments in an aqueous solution are of major importance since only this environment allows the study of native biological processes and mechanical behavior of single molecules in situ. 2. All liquid cells basically perform three functions: contain the sample, contain the liquid, and provide a stable optical path for the laser beam which is reflected off the back of the cantilever. If the optical beam detection method is used, the beam cannot simply pass through a liquid-air interface since it will be refracted all over the place due to movement of the liquid surface. The solution to this problem is to use a glass sighting plate which is submerged in the liquid. 3. Tapping under liquid: There is no capillary force to cause imaging difficulties, so a super stiff cantilever is not required. In practice, the overall oscillation amplitude is set to the smallest value that gives stable imaging (this minimizes the potential damage to the sample) and then the setpoint amplitude-change is increased (increasing the resolution) to the largest value that does not damage the sample.

17 Acoustic Versus Magnetic Drive 1.Cantilevers coated with a magnetic materials can be excited using a nearby electromagnet, known as magnetic ac mode or MACmode TM. The electromagnet is usually a few loops of copper wire wrapped around the liquid cell and driven by a signal generator. 2.Direct bending of the cantilever is induced either as a consequence of magnetostriction or the magnetic interaction of the moment of the film with the applied field. 3.For imaging in fluid, the magnetic ac mode is less noisy than the acoustic drive. 4.For the acoustic drive, significant bending is obtained only near a cantilever bending resonance. In contrast, the magnetic drive yields a broad response that matches the thermal response of the cantilever. The tip image the sample via the medium of an intervening fluid layer. Rapid motion of the molecules in this layer average out any fluid structure, so this layer is not visualized during imaging, but it does affect the overall interaction between tip and sample. Acoustic Magnetic Acoustic Magnetic

18 1. Sample are generally immobile when imaged in air. This situation changes when imaging under liquids. Most biological materials have a high affinity for water and swell or become mobile in its presence. 2. The AFM cantilever operates quite differently in fluid compared to its operation in air or other gasses. In addition to the obvious viscous damping, the cantilever must move fluid with it, leading to a greatly increased effective mass and a substantial reduction of its resonant frequency. 3. The cantilever damping γ scales roughly linearly with the viscosity of the medium and the dimensions of the cantilever. The damping is often parameterized by the mechanical Q-factor. For a simple harmonic oscillator, the damping force, F d, and Q are given by where dd/dt is the cantilever velocity and m is the effective mass of the cantilever. 4. The Q-factor decreases in water and decreases for larger (softer) cantilever. The Q factor in water is about 3 or 4. In air, the same cantilevers would have a Q of >100.

19 Limits on Interaction Forces-Thermal Noise 1. At finite temperature, the tip undergoes thermal motion. At room temperature, the best vertical resolution obtainable with a cantilever with k = 1 N/m is on the order of 0.05 nm. Subangstrom resolution is realized only at low temperatures. 2. The spectral distribution of noise --- harmonic oscillator approximation The Q parameterizes the sharpness of the resonance, being the ratio of the resonance frequency to the full width at half height of the peak. It is also the ratio of the amplitude at resonance to that at a frequency well below resonance. 3. In air or vacuum, high Q can be achieved. There is a large reduction of the thermal noise away from the resonant peak. With operation in water or aqueous buffer, thermal noise is almost equally distributed as a function of frequency because the resonant peak is so broad. 4. The noise spectrum as given by the above two equations forms the basis of a nondestructive method for calibrating cantilever spring constants.

20 Tip-Sample Interactions in Liquids 1. The interactions are strongly dependent on the experimental conditions, such as the tip and sample material, chemical modification of the surfaces, and the surrounding medium. 2. Electrostatic forces are predominant in aqueous solution. They can be mainly attractive or repulsive, depending on the surface potential, the ionic strength, and ph. 3. The use of molecules with appropriate functional groups lead to short-ranging dipole-dipole interactions, hydrogen bonding, or in the case of ligand-receptor couples highly specific complex combinations of different binding types as well as covalent bonding. 4. The interactions between particles in a fluid is a complex many-body problem. 5. It is convenient to group the interactions between tip and sample into the following classes: hydrodynamic, van der Waals, ionic, chemical (e.g. hydrophilic and hydrophobic), and hydration forces. Hydrodynamic Effects Hydrodynamic effect cause interactions between the cantilever and substrate over large distances (comparable to the cantilever dimensions). This is because the free-flow of fluid around the cantilever is disrupted by the presence of a nearby surface, giving rise to an extra damping term R>>D where R is the radius of a sphere a distance D from a flat surface and η is the viscosity of the fluid. This term can dominate the overall damping but it contributes little to image contrast.

21 Tip-Sample Interactions in Liquids Ion and van der Waals interactions 1. Van der Waals forces are weaker in aqueous solution than in a vacuum due to the high dielectric constant of water. Van der Waals forces are of electrodynamic origin and are thus shielded from water dipoles. 2. Colloidal particles aggregate and precipitate as long as van der Waals forces are present. However, dissolved particles carry charges on their surface, which leads to repulsive forces that prevent from aggregation. 3. The electrostatic forces are dependent on the ion concentration and the ph of the solution. In simple term, the ph influences the surface charge, while the number of ions in solution affects the radius of action of the electric field. 4. The theory of Derjaguin, Landau, Verwey, and Overbrek (DLVO) deals with the interplay of electrostatic interactions and short range van der Waals forces. Electrochemical double layer Debye screening length

22 1. The DLVO theory describes the entire interaction as a superposition of the exponential repulsion and the attractive van der Waals force. If the Debye length decreases because of an increase of the ion concentration, the long-range repulsion also decreases and the interaction becomes overall attractive. 2. When nonequally charged surfaces come close together, the interaction is either repulsive or attractive depending on the ratio of the charges. Long-range electrostatic interactions can be monitored and spatially resolved using force spectroscopy. Angew. Chem. Int. Ed. 39, 3212 (2000) Dependence of the force curves on the ion strength for a Si 3 N 4 tip approaching a self-assembled film of 3-sulfanylpropionic acid chemisorbed on gold. Variation of the ion concentration (KCl) Variation of the ph at low ion concentration Numerical simulation Long-range electrostatic repulsion is resolved at low ion concentrations.

23 Hydration (Solvation) Forces 1. If two surfaces approach as close as a couple of nanometers, the forces cannot be sufficiently ascribed with continuum theories, such as attractive van der Waals and repulsive double layer forces. Short-range forces resulting from the molecular structure of the fluid are summarized to the so-called solvation forces. The geometric organization of the solvent molecules at the solid surface plays a crucial role in the strength of these forces. 2. The attractive forces between the solvent molecules and the solid surface lead to an organization of the molecules into quasidiscrete layers. In the case of a fluid enclosed between two solids, one can show that rearrangements are induced as these two surfaces approach, which in most cases provide an exponential decay of an oscillating force. Monotonously declining attractive or repulsive components can also occur, and these are dependent on the geometry of the molecule and the interaction potential with the surface. 3. A special case of the solvation force is the repulsive hydration force observed in aqueous solution. It describes the force necessary to remove the hydration shells of the surface-confined molecules and the bring the tip closer to the surface. Direct experimental observation of solvation forces in an organic solvent at a graphite surface. The figure shows amplitude oscillations as a DFM tip approaches the graphite. The period is equal to the minor axis of the molecule. Angew. Chem. Int. Ed. 39, 3212 (2000)

24 Chemical Force Microscopy (CFM) 1. CFM is done by coating the AFM tip with an ordered monolayer of organic molecules (a selfassembled monolayer) to give it a specific chemical functionality. The force of attraction can be estimated from the excess force required to pull the tip free from the surface. 2. This concept is a particular case of force spectroscopy where the tip and the substrate are derivatized so that interaction between defined functional groups can be clearly distinguished. 3. Functionalization is mostly achieved by covering the tip and sample surface with gold and subsequently chemisorbing thiolated molecules. 4. The environment (atmosphere, electrolyte concentration, ph) plays a central role in CFM. 5. Adhesion forces are generally larger among functional groups that have the ability to form hydrogen bonds than those without. Dependence of the adhesion force F between a tip functionalized with 3- sulfanylpropionic and a similarly prepared substrate on the ph of the solution. Angew. Chem. Int. Ed. 39, 3212 (2000)

25 Force Spectroscopy of Molecular Systems 1. Molecular recognition of biomolecules plays a pivotal role in nature. 2. By functionalization of the AFM tip, it is feasible to measure the forces between two individual molecules locally. 3. Not only static but also dynamic information is available, such as during separation of a single ligand-receptor couple upon applying an external force with different pulling velocities. 4. Long-chain molecules such as DNA, synthetic polymers, polysaccharides, and filamentous proteins experience folding-unfolding cycles upon application of an external force. The characteristic force-extension curves can be utilized to analyze the different conformations of the biomolecules by applying statistic polymer models. 5. If the value of the positive force gradient is the same as the spring constant, a mechanical instability occurs, known as snap-on. The repulsive forces decreases upon retraction of the z- piezo device. The separation from the sample surface occurs at the so-called snap-off. At this point, the maximum attractive force can be measured (adhesion force). Angew. Chem. Int. Ed. 39, 3212 (2000)

26 Van der Waals forces bewteen a passivated Si tip and a Si surface in UHV. Electrostatic interactions between the tip and sample in fluidsattractive forces. Electrostatic interactions between the tip and sample in fluidsrepulsive forces. Specific ligand-receptor interactions between a streptavidin-functionalized tip and a biotinylated surface. Unfolding of a long-chain single molecule. Angew. Chem. Int. Ed. 39, 3212 (2000)

27 Force Spectroscopy of Single Molecules Membrane proteins Imaging individual bacteriorhodopsin molecules in water Separated at a velocity of 40 nm/s while the force spectrum was recorded. Done in buffer solution (300 mm KCl, 10 mm tris-hcl, ph 7.8) Science 288, 143(2000)

28 Model of the 3D structure of BR. (A) BR is a 248- amino acid membrane protein that consists of seven transmembrane α- helices, which are connected by loops. Science 288, 143(2000)

29 Contact-mode image of DNA tightly adsorbed onto a cationic bilayer in buffer. Magnetic DFM of DNA micro-circles on mica in Mgcontaining solution. Microtubule imaged in buffer by magnetic DFM. When katinin is added, the microtubules are digested.

30 A conformational change of E. coli porin OmpF is induced by a ph drop, by a voltage across the sample and by a change of the imaging buffer. Here, the extracellular surface is imaged at (a) ph 7, (b) ph 6, (c) ph2.5, and (d) ph 4. High-speed AFM, combined AFM-optical microscope

31 Locating ligand binding and activation of a single antiporter Mechanical unfolding of a single NhaA molecule. (A) Representative force extension curve recorded on mechanical unfolding of a single NhaA molecule. Each force peak is fitted by the worm-like chain model (red curves), which provides the number of amino-acid (aa) residues stretched. This information was used to assign ends of structural segments (B, red arrows) stabilized by molecular interactions. (B) Secondary structure of NhaA mapped with stable structural segments detected on pulling the carboxyl terminus. Individual structural segments were uniformly shaded grey, whereas the grey gradients reflect uncertainties in determining segment ends. To determine merges of the segments opposite to the atomic force microscopy tip side of the membrane, or within, a membrane thickness of 4 nm was considered. Their corresponding contour lengths are given in brackets. Daniel J Muller EMBO reports 6, 7, 668 (2005)

32 Amyloid Fibrils 1. Amyloid fibrils are highly organized protein aggregates. They were originally discovered as products of uncontrolled protein misfolding, through their involvement in chronic disorders such as Alzheimer s, Parkinson s, and type II diabetes disease. Recently, it has been suggested that amyloidosis could be a universal disease of aging. 2. There is increasing experimental evidence that the amyloid state is a generic protein fold not dependent on amino acid sequence, but often depends on environmental conditions. They are now recognized as a common form of protein structure, in some cases having functional biological roles as bacterial coatings or scaffolds for catalytic reactions. 3. The finding that amyloid fibrils can be formed from a range of very different polypeptide sequences has led to the suggestion that the amyloid configuration is a generic, stable structure of peptides and proteins. This situation contrasts with that of the native states of proteins, where structure is strongly dependent on the specific amino acid sequence, and complex environments or careful regulation is frequently required for correct selfassembly into folded structures.

33 1. Strong correlation between the bending rigidity and the height. 2. The short peptide, comprising the B chain of insulin, assembles into fibrils with a higher modulus than that observed for both morphological forms of the full insulin protein or the other larger proteins. Science 318, 1900 (2007)

34 Adhesive observed around the base of P. linearis holdfasts. Nanotechnology 18, (2007)