A simple method for the determination of qPlus sensor spring constants.

qPlus sensors are widely used to measure forces at the atomic scale, however, confidence in these measurements is limited by inconsistent reports of the spring constant of the sensor and complications from finite tip heights. Here we combine a numerical investigation of the force reconstruction with an experimental characterization of the flexural mechanics of the qPlus sensor. Numerical studies reveal significant errors in reconstructed force for tip heights exceeding 400 µm or one sixth of the cantilever length. Experimental results with a calibrated nanoindenter reveal excellent agreement with an Euler-Bernoulli beam model for the sensor. Prior to the attachment of a tip, measured spring constants of 1902 ± 29 N/m are found to be in agreement with theoretical predictions for the geometry and material properties of the sensor once a peaked ridge in the beam cross section is included. We further develop a correction necessary to adjust the spring constant for the size and placement of the tip.

The impacting cantilever: modal non-convergence and the importance of stiffness matching.

The problem of an Euler-Bernoulli cantilever beam whose free end impacts with a point constraint is revisited from the point of view of modal analysis. It is shown that there is non-uniqueness of consistent impact laws for a given modal truncation. Moreover, taking an N-mode compliant, bilinear formulation and passing to the rigid limit leads to a sequence of impact models that does not converge as N--> ∞. The dynamics of such truncated models are studied numerically and found to give rise to quite different dynamics depending on the number of degrees of freedom taken. The simulations are compared with results from simple experiments that show a propensity for multiple-tap dynamics, in which higher-order modes lead to rapidly cycling intermittent contact. The conclusion reached is that, to derive an accurate model, one needs to avoid the impact limit altogether, and take sufficiently many modes in the formulation to match the actual stiffness of the constraining stop.

High-resolution dynamic atomic force microscopy in liquids with different feedback architectures.

The recent achievement of atomic resolution with dynamic atomic force microscopy (dAFM) [Fukuma et al., Appl. Phys. Lett. 2005, 87, 034101], where quality factors of the oscillating probe are inherently low, challenges some accepted beliefs concerning sensitivity and resolution in dAFM imaging modes. Through analysis and experiment we study the performance metrics for high-resolution imaging with dAFM in liquid media with amplitude modulation (AM), frequency modulation (FM) and drive-amplitude modulation (DAM) imaging modes. We find that while the quality factors of dAFM probes may deviate by several orders of magnitude between vacuum and liquid media, their sensitivity to tip-sample forces can be remarkable similar. Furthermore, the reduction in noncontact forces and quality factors in liquids diminishes the role of feedback control in achieving high-resolution images. The theoretical findings are supported by atomic-resolution images of mica in water acquired with AM, FM and DAM under similar operating conditions.

Drive-amplitude-modulation atomic force microscopy: From vacuum to liquids.

We introduce drive-amplitude-modulation atomic force microscopy as a dynamic mode with outstanding performance in all environments from vacuum to liquids. As with frequency modulation, the new mode follows a feedback scheme with two nested loops: The first keeps the cantilever oscillation amplitude constant by regulating the driving force, and the second uses the driving force as the feedback variable for topography. Additionally, a phase-locked loop can be used as a parallel feedback allowing separation of the conservative and nonconservative interactions. We describe the basis of this mode and present some examples of its performance in three different environments. Drive-amplutide modulation is a very stable, intuitive and easy to use mode that is free of the feedback instability associated with the noncontact-to-contact transition that occurs in the frequency-modulation mode.

Resolving structure and mechanical properties at the nanoscale of viruses with frequency modulation atomic force microscopy.

Structural Biology (SB) techniques are particularly successful in solving virus structures. Taking advantage of the symmetries, a heavy averaging on the data of a large number of specimens, results in an accurate determination of the structure of the sample. However, these techniques do not provide true single molecule information of viruses in physiological conditions. To answer many fundamental questions about the quickly expanding physical virology it is important to develop techniques with the capability to reach nanometer scale resolution on both structure and physical properties of individual molecules in physiological conditions. Atomic force microscopy (AFM) fulfills these requirements providing images of individual virus particles under physiological conditions, along with the characterization of a variety of properties including local adhesion and elasticity. Using conventional AFM modes is easy to obtain molecular resolved images on flat samples, such as the purple membrane, or large viruses as the Giant Mimivirus. On the contrary, small virus particles (25-50 nm) cannot be easily imaged. In this work we present Frequency Modulation atomic force microscopy (FM-AFM) working in physiological conditions as an accurate and powerful technique to study virus particles. Our interpretation of the so called "dissipation channel" in terms of mechanical properties allows us to provide maps where the local stiffness of the virus particles are resolved with nanometer resolution. FM-AFM can be considered as a non invasive technique since, as we demonstrate in our experiments, we are able to sense forces down to 20 pN. The methodology reported here is of general interest since it can be applied to a large number of biological samples. In particular, the importance of mechanical interactions is a hot topic in different aspects of biotechnology ranging from protein folding to stem cells differentiation where conventional AFM modes are already being used.

Gaining insight into the physics of dynamic atomic force microscopy in complex environments using the VEDA simulator.

Dynamic atomic force microscopy (dAFM) continues to grow in popularity among scientists in many different fields, and research on new methods and operating modes continues to expand the resolution, capabilities, and types of samples that can be studied. But many promising increases in capability are accompanied by increases in complexity. Indeed, interpreting modern dAFM data can be challenging, especially on complicated material systems, or in liquid environments where the behavior is often contrary to what is known in air or vacuum environments. Mathematical simulations have proven to be an effective tool in providing physical insight into these non-intuitive systems. In this article we describe recent developments in the VEDA (virtual environment for dynamic AFM) simulator, which is a suite of freely available, open-source simulation tools that are delivered through the cloud computing cyber-infrastructure of nanoHUB (www.nanohub.org). Here we describe three major developments. First, simulations in liquid environments are improved by enhancements in the modeling of cantilever dynamics, excitation methods, and solvation shell forces. Second, VEDA is now able to simulate many new advanced modes of operation (bimodal, phase-modulation, frequency-modulation, etc.). Finally, nineteen different tip-sample models are available to simulate the surface physics of a wide variety different material systems including capillary, specific adhesion, van der Waals, electrostatic, viscoelasticity, and hydration forces. These features are demonstrated through example simulations and validated against experimental data, in order to provide insight into practical problems in dynamic AFM.

Calibration of higher eigenmode spring constants of atomic force microscope cantilevers.

Standard spring constant calibration methods are compared when applied to higher eigenmodes of cantilevers used in dynamic atomic force microscopy (dAFM). Analysis shows that Sader's original method (Sader et al 1999 Rev. Sci. Instrum. 70 3967-9), which relies on a priori knowledge of the eigenmode shape, is poorly suited for the calibration of higher eigenmodes. On the other hand, the thermal noise method (Hutter and Bechhoefer 1993 Rev. Sci. Instrum. 64 1868-73) does not require knowledge of the eigenmode and remains valid for higher eigenmodes of the dAFM probe. Experimental measurements of thermal vibrations in air for three representative cantilevers are provided to support the theoretical results.

Origins of phase contrast in the atomic force microscope in liquids.

We study the physical origins of phase contrast in dynamic atomic force microscopy (dAFM) in liquids where low-stiffness microcantilever probes are often used for nanoscale imaging of soft biological samples with gentle forces. Under these conditions, we show that the phase contrast derives primarily from a unique energy flow channel that opens up in liquids due to the momentary excitation of higher eigenmodes. Contrary to the common assumption, phase-contrast images in liquids using soft microcantilevers are often maps of short-range conservative interactions, such as local elastic response, rather than tip-sample dissipation. The theory is used to demonstrate variations in local elasticity of purple membrane and bacteriophage 29 virions in buffer solutions using the phase-contrast images.

Compositional contrast of biological materials in liquids using the momentary excitation of higher eigenmodes in dynamic atomic force microscopy.

Atomic Force microscope (AFM) cantilevers commonly used for imaging soft biological samples in liquids experience a momentary excitation of the higher eigenmodes at each tap. This transient response is very sensitive to the local sample elasticity under gentle imaging conditions because the higher eigenmode time period is comparable to the tip-sample contact time. By mapping the momentary excitation response, we demonstrate a new scanning probe spectroscopy capable of resolving with high sensitivity the variations in the elasticity of soft biological materials in liquids.

VEDA: a web-based virtual environment for dynamic atomic force microscopy.

We describe here the theory and applications of virtual environment dynamic atomic force microscopy (VEDA), a suite of state-of-the-art simulation tools deployed on nanoHUB (www.nanohub.org) for the accurate simulation of tip motion in dynamic atomic force microscopy (dAFM) over organic and inorganic samples. VEDA takes advantage of nanoHUB's cyberinfrastructure to run high-fidelity dAFM tip dynamics computations on local clusters and the teragrid. Consequently, these tools are freely accessible and the dAFM simulations are run using standard web-based browsers without requiring additional software. A wide range of issues in dAFM ranging from optimal probe choice, probe stability, and tip-sample interaction forces, power dissipation, to material property extraction and scanning dynamics over hetereogeneous samples can be addressed.