► Development of small cantilever. ► Speed increase by a factor of ten using small cantilevers on a commercial AFM. ► Force sensitivity increase by a factor of five using small cantilever prototypes for force spectroscopy measurements.
In this study, we demonstrate the increased performance in speed and sensitivity achieved by the use of small AFM cantilevers on a standard AFM system. For this, small rectangular silicon oxynitride cantilevers were utilized to arrive at faster atomic force microscopy (AFM) imaging times and more sensitive molecular recognition force spectroscopy (MRFS) experiments. The cantilevers we used had lengths between 13 and 46 μm, a width of about 11 μm, and a thickness between 150 and 600 nm. They were coated with chromium and gold on the backside for a better laser reflection. We characterized these small cantilevers through their frequency spectrum and with electron microscopy. Due to their small size and high resonance frequency we were able to increase the imaging speed by a factor of 10 without any loss in resolution for images from several μm scansize down to the nanometer scale. This was shown on bacterial surface layers (s-layer) with tapping mode under aqueous, near physiological conditions and on nuclear membranes in contact mode in ambient environment. In addition, we showed that single molecular forces can be measured with an up to 5 times higher force sensitivity in comparison to conventional cantilevers with similar spring constants.
Small cantilever; High resolution imaging; Fast AFM imaging; Ultra-sensitive molecular recognition force spectroscopy
Microcantilevers were first introduced as imaging probes in Atomic Force Microscopy (AFM) due to their extremely high sensitivity in measuring surface forces. The versatility of these probes, however, allows the sensing and measurement of a host of mechanical properties of various materials. Sensor parameters such as resonance frequency, quality factor, amplitude of vibration and bending due to a differential stress can all be simultaneously determined for a cantilever. When measuring the mechanical properties of materials, identifying and discerning the most influential parameters responsible for the observed changes in the cantilever response are important. We will, therefore, discuss the effects of various force fields such as those induced by mass loading, residual stress, internal friction of the material, and other changes in the mechanical properties of the microcantilevers. Methods to measure variations in temperature, pressure, or molecular adsorption of water molecules are also discussed. Often these effects occur simultaneously, increasing the number of parameters that need to be concurrently measured to ensure the reliability of the sensors. We therefore systematically investigate the geometric and environmental effects on cantilever measurements including the chemical nature of the underlying interactions. To address the geometric effects we have considered cantilevers with a rectangular or circular cross section. The chemical nature is addressed by using cantilevers fabricated with metals and/or dielectrics. Selective chemical etching, swelling or changes in Young's modulus of the surface were investigated by means of polymeric and inorganic coatings. Finally to address the effect of the environment in which the cantilever operates, the Knudsen number was determined to characterize the molecule-cantilever collisions. Also bimaterial cantilevers with high thermal sensitivity were used to discern the effect of temperature variations. When appropriate, we use continuum mechanics, which is justified according to the ratio between the cantilever thickness and the grain size of the materials. We will also address other potential applications such as the ageing process of nuclear materials, building materials, and optical fibers, which can be investigated by monitoring their mechanical changes with time. In summary, by virtue of the dynamic response of a miniaturized cantilever shaped material, we present useful measurements of the associated elastic properties.
Microcantilever; mechanics; ageing; environment; stress; gas; materials; sensor; pressure; temperature
We critically discuss the extraction of intrinsic cantilever properties, namely eigenfrequency f
n, quality factor Q
n and specifically the stiffness k
n of the nth cantilever oscillation mode from thermal noise by an analysis of the power spectral density of displacement fluctuations of the cantilever in contact with a thermal bath. The practical applicability of this approach is demonstrated for several cantilevers with eigenfrequencies ranging from 50 kHz to 2 MHz. As such an analysis requires a sophisticated spectral analysis, we introduce a new method to determine k
n from a spectral analysis of the demodulated oscillation signal of the excited cantilever that can be performed in the frequency range of 10 Hz to 1 kHz regardless of the eigenfrequency of the cantilever. We demonstrate that the latter method is in particular useful for noncontact atomic force microscopy (NC-AFM) where the required simple instrumentation for spectral analysis is available in most experimental systems.
AFM; cantilever; noncontact atomic force microscopy (NC-AFM); Q-factor; thermal excitation; resonance; spectral analysis; stiffness
The noise of the frequency-shift signal Δf in noncontact atomic force microscopy (NC-AFM) consists of cantilever thermal noise, tip–surface-interaction noise and instrumental noise from the detection and signal processing systems. We investigate how the displacement-noise spectral density d
z at the input of the frequency demodulator propagates to the frequency-shift-noise spectral density d
f at the demodulator output in dependence of cantilever properties and settings of the signal processing electronics in the limit of a negligible tip–surface interaction and a measurement under ultrahigh-vacuum conditions. For a quantification of the noise figures, we calibrate the cantilever displacement signal and determine the transfer function of the signal-processing electronics. From the transfer function and the measured d
z, we predict d
f for specific filter settings, a given level of detection-system noise spectral density d
ds and the cantilever-thermal-noise spectral density d
th. We find an excellent agreement between the calculated and measured values for d
f. Furthermore, we demonstrate that thermal noise in d
f, defining the ultimate limit in NC-AFM signal detection, can be kept low by a proper choice of the cantilever whereby its Q-factor should be given most attention. A system with a low-noise signal detection and a suitable cantilever, operated with appropriate filter and feedback-loop settings allows room temperature NC-AFM measurements at a low thermal-noise limit with a significant bandwidth.
Cantilever; feedback loop; filter; noncontact atomic force microscopy (NC-AFM); noise
Micro-cantilever arrays with different dimensions are fabricated by micromachining technique onto silicon <1 0 0> substrate. These sputtered Gold-Coated micro-cantilevers were later surface functionalized. Scanning Electron Microscopy, Atomic Force Microscopy and Optical SWLI using LASER probe are employed to characterize the morphology and image measurement of the micro-cantilever arrays, respectively. Compared with conventional AFM and SPM measurement technique, the proposed method has demonstrated sufficient flexibility and reliability. The experimental results have been analyzed and presented in this paper for MEMS Micro-cantilevers. The scanning White Light Interferometry based two point high resolution optical method is presented for characterizing Micro-cantilevers and other MEMS micro-structures. The repeatable error and the repeatable precision produced in the proposed image measurement method is nanometre confirmable. In this piece of work, we investigate the micro-structure fabrication and image measurement of Length, Width and Step-Height of micro-cantilever arrays fabricated using bulk micromachining technique onto Silicon <100> substrate.
Scanning Electron Microscopy; Atomic Force Microscopy; Micro-cantilever; Optics; Image Measurement; Silicon (100), Scanning White Light Interferometry
Intermodulation atomic force microscopy (ImAFM) is a mode of dynamic atomic force microscopy that probes the nonlinear tip–surface force by measurement of the mixing of multiple modes in a frequency comb. A high-quality factor cantilever resonance and a suitable drive comb will result in tip motion described by a narrow-band frequency comb. We show, by a separation of time scales, that such motion is equivalent to rapid oscillations at the cantilever resonance with a slow amplitude and phase or frequency modulation. With this time-domain perspective, we analyze single oscillation cycles in ImAFM to extract the Fourier components of the tip–surface force that are in-phase with the tip motion (F
I) and quadrature to the motion (F
Q). Traditionally, these force components have been considered as a function of the static-probe height only. Here we show that F
I and F
Q actually depend on both static-probe height and oscillation amplitude. We demonstrate on simulated data how to reconstruct the amplitude dependence of F
I and F
Q from a single ImAFM measurement. Furthermore, we introduce ImAFM approach measurements with which we reconstruct the full amplitude and probe-height dependence of the force components F
I and F
Q, providing deeper insight into the tip–surface interaction. We demonstrate the capabilities of ImAFM approach measurements on a polystyrene polymer surface.
atomic force microscopy; AFM; frequency combs; force spectroscopy; high-quality-factor resonators; intermodulation; multifrequency
Atomic force microscopy (AFM) has become a powerful tool for measuring material properties in biology and imposing mechanical boundary conditions on samples from single molecules to cells and tissues. Constant force or constant height can be maintained in an AFM experiment through feedback control of cantilever deflection, known respectively as a ‘force clamp’ or ‘position clamp’. However, stiffness, the third variable in the Hookean relation F = kx that describes AFM cantilever deflection, has not been dynamically controllable in the same way. Here we present and demonstrate a ‘stiffness clamp’ that can vary the apparent stiffness of an AFM cantilever. This method, employable on any AFM system by modifying feedback control of the cantilever, allows rapid and reversible tuning of the stiffness exposed to the sample in a way that can decouple the role of stiffness from force and deformation. We demonstrated the AFM stiffness clamp on two different samples: a contracting fibroblast cell and an expanding polyacrylamide hydrogel. We found that the fibroblast, a cell type that secretes and organizes the extracellular matrix, exhibited a rapid, sub-second change in traction rate (dF/dt) and contraction velocity (dx/dt) in response to step changes in stiffness between 1–100 nN/µm. This response was independent of the absolute contractile force and cell height, demonstrating that cells can react directly to changes in stiffness alone. In contrast, the hydrogel used in our experiment maintained a constant expansion velocity (dx/dt) over this range of stiffness, while the traction rate (dF/dt) changed with stiffness, showing that passive materials can also behave differently in different stiffness environments. The AFM stiffness clamp presented here, which is applicable to mechanical measurements on both biological and non-biological samples, may be used to investigate cellular mechanotransduction under a wide range of controlled mechanical boundary conditions.
Atomic force microscopy (AFM) allows for the visualizing of individual proteins, DNA
molecules, protein-protein complexes, and DNA-protein complexes. On the end of the
microscope's cantilever is a nano-scale probe, which traverses image areas ranging from
nanometers to micrometers, measuring the elevation of macromolecules resting on the
substrate surface at any given point. Electrostatic forces cause proteins, lipids, and
nucleic acids to loosely attach to the substrate in random orientations and permit
imaging. The generated data resemble a topographical map, where the macromolecules resolve
as three-dimensional particles of discrete sizes (Figure 1) 1,2.
Tapping mode AFM involves the repeated oscillation of the cantilever, which permits
imaging of relatively soft biomaterials such as DNA and proteins. One of the notable
benefits of AFM over other nanoscale microscopy techniques is its relative adaptability to
visualize individual proteins and macromolecular complexes in aqueous buffers, including
near-physiologic buffered conditions, in real-time, and without staining or coating the
sample to be imaged.
The method presented here describes the imaging of DNA and an immunoadsorbed
transcription factor (i.e. the glucocorticoid receptor, GR) in buffered solution
(Figure 2). Immunoadsorbed proteins and protein complexes can be separated
from the immunoadsorbing antibody-bead pellet by competition with the antibody epitope and
then imaged (Figure 2A). This allows for biochemical manipulation of the
biomolecules of interest prior to imaging. Once purified, DNA and proteins can be mixed
and the resultant interacting complex can be imaged as well. Binding of DNA to mica
requires a divalent cation 3,such as Ni2+ or Mg2+, which
can be added to sample buffers yet maintain protein activity. Using a similar approach,
AFM has been utilized to visualize individual enzymes, including RNA polymerase
4 and a repair enzyme 5, bound to individual DNA strands. These
experiments provide significant insight into the protein-protein and DNA-protein
biophysical interactions taking place at the molecular level. Imaging individual
macromolecular particles with AFM can be useful for determining particle homogeneity and
for identifying the physical arrangement of constituent components of the imaged
particles. While the present method was developed for visualization of GR-chaperone
protein complexes 1,2 and DNA strands to which the GR can bind, it can be
applied broadly to imaging DNA and protein samples from a variety of sources.
The size of a cell is a fundamental physiological property and is closely regulated by various environmental and genetic factors. Optical or confocal microscopy can be used to measure the dimensions of adherent cells, and Coulter counter or flow cytometry (forward scattering light intensity) can be used to estimate the volume of single cells in a flow. Although these methods could be used to obtain the mass of single live cells, no method suitable for directly measuring the mass of single adherent cells without detaching them from the surface is currently available. We report the design, fabrication, and testing of ‘living cantilever arrays’, an approach to measure the mass of single adherent live cells in fluid using silicon cantilever mass sensor. HeLa cells were injected into microfluidic channels with a linear array of functionalized silicon cantilevers and the cells were subsequently captured on the cantilevers with positive dielectrophoresis. The captured cells were then cultured on the cantilevers in a microfluidic environment and the resonant frequencies of the cantilevers were measured. The mass of a single HeLa cell was extracted from the resonance frequency shift of the cantilever and was found to be close to the mass value calculated from the cell density from the literature and the cell volume obtained from confocal microscopy. This approach can provide a new method for mass measurement of a single adherent cell in its physiological condition in a non-invasive manner, as well as optical observations of the same cell. We believe this technology would be very valuable for single cell time-course studies of adherent live cells.
Detection of magnetic resonance as a force between a magnetic tip and nuclear spins has previously been shown to enable sub-10 nm resolution 1H imaging. Maximizing the spin force in such a magnetic resonance force microscopy (MRFM) experiment demands a high field gradient. In order to study a wide range of samples, it is equally desirable to locate the magnetic tip on the force sensor. Here we report the development of attonewton-sensitivity cantilevers with high gradient cobalt nanomagnet tips. The damage layer thickness and saturation magnetization of the magnetic material were characterized by X-ray photoelectron spectroscopy and superconducting quantum interference device magnetometry. The coercive field and saturation magnetization of an individual tip were quantified in situ using frequency-shift cantilever magnetometry. Measurements of cantilever dissipation versus magnetic field and tip-sample separation were conducted. MRFM signals from protons in a polystyrene film were studied versus rf irradiation frequency and tip-sample separation, and from this data the tip field and tip-field gradient were evaluated. Magnetic tip performance was assessed by numerically modeling the frequency dependence of the magnetic resonance signal. We observed a tip-field gradient ∂Bztip∕∂z estimated to be between 4.4 and 5.4 MT m−1, which is comparable to the gradient used in recent 4 nm resolution 1H imaging experiments and larger by nearly an order of magnitude than the gradient achieved in prior magnet-on-cantilever MRFM experiments.
nanofabrication; magnetic resonance force microscopy; magnetometry; X-ray photoelectron spectroscopy; superconducting quantum interference device; surface-induced dissipation
This study presents the deflection, resonant frequency and stress results of rectangular, triangular, and step profile microcantilevers subject to surface stress. These cantilevers can be used as the sensing element in microcantilever biosensors. To increase the overall sensitivity of microcantilever biosensors, both the deflection and the resonant frequency of the cantilever should be increased. The effect of the cantilever profile change and the cantilever cross-section shape change is first investigated separately and then together. A finite element code ANSYS Multiphysics is used and solid finite elements cantilever models are solved. A surface stress of 0.05 N/m was applied to the top surface of the cantilevers. The cantilevers are made of silicon with elastic modulus 130 GPa and Poisson’s ratio 0.28. To show the conformity of this study, the numerical results are compared against their analytical ones. Results show that triangular and step cantilevers have better deflection and frequency characteristics than rectangular ones.
biosensor; surface stress; microcantilever; resonant frequency; deflection
To demonstrate the feasibility of measuring the elasticity of intact crystalline lenses using atomic force microscopy (AFM).
AFM elasticity measurements were performed on intact lenses from 18 fresh cynomolgus monkey cadaver eyes (4-10 years old, <1 day postmortem) that had been left attached to their zonule-ciliary body-sclera framework. The eyes were prepared by bonding a plastic ring on the sclera after removal of the conjunctival, adipose, and muscle tissues. The posterior pole was sectioned, with the excess vitreous removed, and the eye's anterior section was placed on a Teflon slide to protect the posterior pole of the lens. The cornea and iris were then sectioned. The lens-zonule-ciliary body-sclera section was then placed in a Petri dish filled with balanced salt solution in an AFM system designed for force measurements. Next, the central pole of the anterior surface of the intact lens was probed with the AFM cantilever tip. The recorded AFM cantilever deflection-indentation curves were used to derive force-indentation curves for the lens after factoring out the deflection of the cantilever on a hard surface. Young's modulus of the lens was calculated from the force-indentation relation using the Hertz model.
Young's modulus was 1,720±880 Pa (range: 409-3,210 Pa) in the 18 cynomolgus monkey lenses.
AFM can be used to provide measurements of the elasticity of the whole lens including the capsule. Values obtained using AFM on cynomolgus monkey lenses are similar to published values obtained using dynamic mechanical analysis on young human lenses.
One of the bio-sensing mechanisms is mechanical. Rather than measuring shift in resonance frequency, we adopt to measure the change in spring constant due to adsorption, as one of the fundamental sensing mechanism. This study explains determination of spring constant of a surface functionalized micro machined micro cantilever, which resonates in a trapezoidal cavity-on Silicon <100> wafer, with the resonating frequency of 7000 cycles per second. This thin-flimsy-oxide micro-cantilever has a typical shape, and the tip of the micro-cantilever is dip-coated with chemically and biologically active material. The change in mass, due to adsorption, is detected by measuring the change in spring constant. The Force-Distance spectroscopy is used to detect the change in spring constant. The experimental results, show that the mechanical sensing scheme used, permit this surface functionalized micro machined micro cantilever to be used as a molecular mass sensor. The mechanical spring behaviour of a micro-cantilever, a micro-mechanical device can be used to develop ultra-tech micro-mechanical system using computer interface.
Micromachining; Micro-cantilever; Silicon<100>; Bio-Sensor; F-d Spectroscopy
We present a method for characterizing ultrathin films using sensitivity-enhanced atomic force acoustic microscopy, where a concentrated-mass cantilever having a flat tip was used as a sensitive oscillator. Evaluation was aimed at 6-nm-thick and 10-nm-thick diamond-like carbon (DLC) films deposited, using different methods, on a hard disk for the effective Young's modulus defined as E/(1 - ν2), where E is the Young's modulus, and ν is the Poisson's ratio. The resonant frequency of the cantilever was affected not only by the film's elasticity but also by the substrate even at an indentation depth of about 0.6 nm. The substrate effect was removed by employing a theoretical formula on the indentation of a layered half-space, together with a hard disk without DLC coating. The moduli of the 6-nm-thick and 10-nm-thick DLC films were 392 and 345 GPa, respectively. The error analysis showed the standard deviation less than 5% in the moduli.
Atomic force acoustic microscopy; Thin film; Elastic modulus; Diamond-like carbon; Concentrated-mass cantilever
Atomic force microscope (AFM) probe with a long and rigid needle tip was fabricated and studied for high Q factor dynamic (tapping mode) AFM imaging of samples submersed in liquid. The extended needle tip over a regular commercially-available tapping mode AFM cantilever was sufficiently long to keep the AFM cantilever from submersed in liquid, which significantly minimized the hydrodynamic damping involved in dynamic AFM imaging of samples in liquid. Dynamic AFM imaging of samples in liquid at an intrinsic Q factor of over 100 and an operation frequency of over 200 kHz was demonstrated. The method has the potential to be extended to acquire viscoelastic materials properties and provide truly gentle imaging of soft biological samples in physiological environments.
Efficient maintenance of glucose homeostasis is a major challenge in diabetes therapy, where accurate and reliable glucose level detection is required. Though several methods are currently used, these suffer from impaired response and often unpredictable drift, making them unsuitable for long-term therapeutic practice. In this study, we demonstrate a method that uses a functionalized atomic force microscope (AFM) cantilever as the sensor for reliable glucose detection with sufficient sensitivity and selectivity for clinical use. We first modified the AFM tip with aminopropylsilatrane (APS) and then adsorbed glucose-specific lectin concanavalin A (Con A) onto the surface. The Con A/APS-modified probes were then used to detect glucose by monitoring shifts in the cantilever resonance frequency. To confirm the molecule-specific interaction, AFM topographical images were acquired of identically treated silicon substrates which indicated a specific attachment for glucose-Con A and not for galactose-Con A. These results demonstrate that by monitoring the frequency shift of the AFM cantilever, this sensing system can detect the interaction between Con A and glucose, one of the biomolecule recognition processes, and may assist in the detection and mass quantification of glucose for clinical applications with very high sensitivity.
Atomic force microscopy (AFM) in contact mode and tapping mode is employed for high resolution studies of soft organic molecules (fetal bovine serum proteins) on hard inorganic diamond substrates in solution and air. Various effects in morphology and phase measurements related to the cantilever spring constant, amplitude of tip oscillations, surface approach, tip shape and condition are demonstrated and discussed based on the proposed schematic models. We show that both diamond and proteins can be mechanically modified by Si AFM cantilever. We propose how to choose suitable cantilever type, optimize scanning parameters, recognize and minimize various artifacts, and obtain reliable AFM data both in solution and in air to reveal microscopic characteristics of protein-diamond interfaces. We also suggest that monocrystalline diamond is well defined substrate that can be applicable for fundamental studies of molecules on surfaces in general.
Microcantilevers are used in a number of applications including atomic-force microscopy (AFM). In this work, deflection-sensing elements along with heating elements are integrated onto micromachined cantilever arrays to increase sensitivity, and reduce complexity and cost. An array of probes with 5–10 nm gold ultrathin film sensors on silicon substrates for high throughput scanning probe microscopy is developed. The deflection sensitivity is 0.2 ppm/nm. Plots of the change in resistance of the sensing element with displacement are used to calibrate the probes and determine probe contact with the substrate. Topographical scans demonstrate high throughput and nanometer resolution. The heating elements are calibrated and the thermal coefficient of resistance (TCR) is 655 ppm/K. The melting temperature of a material is measured by locally heating the material with the heating element of the cantilever while monitoring the bending with the deflection sensing element. The melting point value measured with this method is in close agreement with the reported value in literature.
Microcantilevers; Scanning probe microscopy; Piezoresistive sensing; Parallel imaging; Elastography; Mechanical characterization; Melting point; High throughput
We studied nanoscale mechanical properties of PC12 living cells with a Force Feedback Microscope using two experimental approaches. The first one consists in measuring the local mechanical impedance of the cell membrane while simultaneously mapping the cell morphology at constant force. As the interaction force is increased, we observe the appearance of the sub-membrane cytoskeleton. We compare our findings with the outcome of other techniques. The second experimental approach consists in a spectroscopic investigation of the cell while varying the tip indentation into the membrane and consequently the applied force. At variance with conventional dynamic Atomic Force Microscopy techniques, here it is not mandatory to work at the first oscillation eigenmode of the cantilever: the excitation frequency of the tip can be chosen arbitrary leading then to new spectroscopic AFM techniques. We found in this way that the mechanical response of the PC12 cell membrane is found to be frequency dependent in the 1 kHz - 10 kHz range. In particular, we observe that the damping coefficient consistently decreases when the excitation frequency is increased.
We study the coupling of lateral and normal tip oscillations and its effect on the imaging process of frequency-modulated dynamic atomic force microscopy. The coupling is induced by the interaction between tip and surface. Energy is transferred from the normal to the lateral excitation, which can be detected as damping of the cantilever oscillation. However, energy can be transferred back into the normal oscillation, if not dissipated by the usually uncontrolled mechanical damping of the lateral excitation. For certain cantilevers, this dissipation mechanism can lead to dissipation rates larger than 0.01 eV per period. The mechanism produces an atomic contrast for ionic crystals with two maxima per unit cell in a line scan.
atomic force microscopy (AFM); frequency-modulated atomic force microscopy (FM-AFM); energy dissipation
Atomic force microscopy (AFM) is widely used in the biological sciences. Despite 25 years of technical developments, two popular modes of bioAFM, imaging and single molecule force spectroscopy, remain hindered by relatively poor force precision and stability. Recently, we achieved both sub-pN force precision and stability under biologically useful conditions (in liquid at room temperature). Importantly, this sub-pN level of performance is routinely accessible using a commercial cantilever on a commercial instrument. The two critical results are that (i) force precision and stability were limited by the gold coating on the cantilevers, and (ii) smaller yet stiffer cantilevers did not lead to better force precision on time scales longer than 25 ms. These new findings complement our previous work that addressed tip-sample stability. In this review, we detail the methods needed to achieve this sub-pN force stability and demonstrate improvements in force spectroscopy and imaging when using uncoated cantilevers. With this improved cantilever performance, the widespread use of nonspecific biomolecular attachments becomes a limiting factor in high-precision studies. Thus, we conclude by briefly reviewing site-specific covalent-immobilization protocols for linking a biomolecule to the substrate and to the AFM tip.
Atomic force microscopy; scanning probe microscopy; single molecule force spectroscopy; drift; overstretching DNA; cantilever; imaging
Biological membranes contain ion channels, which are nanoscale pores allowing controlled ionic transport and mediating key biological functions underlying normal/abnormal living. Synthetic membranes with defined pores are being developed to control various processes, including filtration of pollutants, charge transport for energy storage, and separation of fluids and molecules. Although ionic transport (currents) can be measured with single channel resolution, imaging their structure and ionic currents simultaneously is difficult. Atomic force microscopy enables high resolution imaging of nanoscale structures and can be modified to measure ionic currents simultaneously. Moreover, the ionic currents can also be used to image structures. A simple method for fabricating conducting AFM cantilevers to image pore structures at high resolution is reported. Tungsten microwires with nanoscale tips are insulated except at the apex. This allows simultaneous imaging via cantilever deflections in normal AFM force feedback mode as well as measuring localized ionic currents. These novel probes measure ionic currents as small as picoampere while providing nanoscale spatial resolution surface topography and is suitable for measuring ionic currents and conductance of biological ion channels.
The atomic force microscope was used to image adsorption of a monoclonal IgM on mica in real time. Under the smallest possible force we could achieve (<4 nN), the cantilever tip behaved as a molecular broom and was observed to orient protein aggregates in strands oriented perpendicularly to the facet of the cantilever tip. Rotating the scan direction preserved the orientational relationship, as seen by the formation of rotated strands. When the applied force was increased, the distance between the strands increased, indicating the amount of protein that can be swept depends on the applied force. The effect of scanning increased the apparent surface coverage of IgM. Manipulation of a deposited fibrinogen layer with a 4-nN repulsive force was observed only after tens of minutes, but not to the extent that strands formed, indicating a greater adhesion between the fibrinogen and mica than between IgM and mica. With an applied repulsive force of 30 nN, fibrinogen strands formed and the protein was manipulated to produce the block letter U. At a much higher repulsive force, the entire scanning area was swept clean.
The volume required for the rheological characterization of fluids can be minimized by using micromechanical cantilevers as viscosity sensors. Here, a simple measurement tool for the characterization of sugar solutions is proposed. The sensor consists of a micromechanical cantilever as used in an atomic force microscopy which is integrated into a closed fluid handling system. Fluid properties are derived from an analysis of the power spectral density of the fluctuations of the cantilever deflection signal. The data acquisition system is operated with standard consumer computer components, which limits the costs for the hardware. Measurements with different sugar solutions indicate that the sensor system provides reliable viscosity values for sugar concentrations as they occur in biological systems. The viscosities of the sugar solutions could be evaluated with an error smaller than 5 %.
cantilever sensor; viscosity; sugar solution; thermal noise
The resonance frequency, amplitude, and phase response of the first two eigenmodes of two contact-resonance atomic force microscopy (CR-AFM) configurations, which differ in the method used to excite the system (cantilever base vs sample excitation), are analyzed in this work. Similarities and differences in the observables of the cantilever dynamics, as well as the different effect of the tip–sample contact properties on those observables in each configuration are discussed. Finally, the expected accuracy of CR-AFM using phase-locked loop detection is investigated and quantification of the typical errors incurred during measurements is provided.
contact-resonance AFM; dynamic AFM; frequency modulation; phase-locked loop; viscoelasticity