Frequency Modulation Mode Research Articles

Single Molecular Observation of DNA and DNA Complexes by Atomic Force Microscopy

Atomic force microscopy (AFM) provides a novel way to understand the structure-function relationship of protein synthesis at a single-molecular level. High-resolution AFM imaging in air, liquid and vacuum allows for single DNA, RNA and proteins to be observed at the nano-scale in addition to their conformational transitions, bound states, temporal behavior, and assembly. The recent development of frequency modulation mode AFM has led to imaging hydration structures of DNA in water and molecular polarization of DNA complexes in vacuum. Real-time imaging in near-physiological environments captures transcriptional activation with characteristic conformation of DNA-protein complexes. We review current achievements and the future potential of methodological and biological AFM to reveal insights into DNA, RNA and their complexes.

Multiscale approach for simulations of Kelvin probe force microscopy with atomic resolution

The distance dependence and atomic-scale contrast recently observed in nominal contact potential difference (CPD) signals simultaneously recorded by Kelvin probe force microscopy (KPFM) using noncontact atomic force microscopy (NCAFM) on defect-free surfaces of insulating as well as semiconducting samples have stimulated theoretical attempts to explain such effects. Especially in the case of insulators, it is not quite clear how the applied bias voltage affects electrostatic forces acting on the atomic scale. We attack this problem in two steps. First, the electrostatics of the macroscopic tip-cantilever-sample system is treated by a finite-difference method on an adjustable nonuniform mesh. Then the resulting electric field under the tip apex is inserted into a series of atomistic wavelet-based density functional theory (DFT) calculations. Results are shown for a realistic neutral but reactive silicon nanoscale tip interacting with a NaCl(001) sample. Bias-dependent forces and resulting atomic displacements are computed to within an unprecedented accuracy. Theoretical expressions for amplitude modulation (AM) and frequency modulation (FM) KPFM signals and for the corresponding local contact potential differences (LCPD) are obtained by combining the macroscopic and atomistic contributions to the electrostatic force component generated at the voltage modulation frequency, and evaluated for several tip oscillation amplitudes $A$ up to 10 nm. For $A$ $=$ 0.1 \AA{}, the computed LCPD contrast is proportional to the slope of the atomistic force versus bias in the AM mode and to its derivative with respect to the tip-sample separation in the FM mode. Being essentially constant over a few volts, this slope is the basic quantity that determines variations of the atomic-scale LCPD contrast. Already above $A$ $=$ 1 \AA{}, the LCPD contrasts in both modes exhibit almost the same spatial dependence as the slope. In the AM mode, this contrast is approximately proportional to ${A}^{\ensuremath{-}1/2}$, but remains much weaker than the contrast in the FM mode, which drops somewhat faster as $A$ is increased. These trends are a consequence of the macroscopic contributions to the KPFM signal, which are stronger in the AM-mode and especially important if the sample is an insulator even at subnanometer separations where atomic-scale contrast appears.

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.

QPlus magnetic force microscopy in frequency-modulation mode with millihertz resolution

Magnetic force microscopy (MFM) allows one to image the domain structure of ferromagnetic samples by probing the dipole forces between a magnetic probe tip and a magnetic sample. The magnetic domain structure of the sample depends on the alignment of the individual atomic magnetic moments. It is desirable to be able to image both individual atoms and domain structures with a single probe. However, the force gradients of the interactions responsible for atomic contrast and those causing domain contrast are orders of magnitude apart, ranging from up to 100 Nm−1 for atomic interactions down to 0.0001 Nm−1 for magnetic dipole interactions. Here, we show that this gap can be bridged with a qPlus sensor, with a stiffness of 1800 Nm−1 (optimized for atomic interaction), which is sensitive enough to measure millihertz frequency contrast caused by magnetic dipole–dipole interactions. Thus we have succeeded in establishing a sensing technique that performs scanning tunneling microscopy, atomic force microscopy and MFM with a single probe.

Development of a Hybrid Atomic Force Microscopic Measurement System Combined with White Light Scanning Interferometry

A hybrid atomic force microscopic (AFM) measurement system combined with white light scanning interferometry for micro/nanometer dimensional measurement is developed. The system is based on a high precision large-range positioning platform with nanometer accuracy on which a white light scanning interferometric module and an AFM head are built. A compact AFM head is developed using a self-sensing tuning fork probe. The head need no external optical sensors to detect the deflection of the cantilever, which saves room on the head, and it can be directly fixed under an optical microscopic interferometric system. To enhance the system’s dynamic response, the frequency modulation (FM) mode is adopted for the AFM head. The measuring data can be traceable through three laser interferometers in the system. The lateral scanning range can reach 25 mm × 25 mm by using a large-range positioning platform. A hybrid method combining AFM and white light scanning interferometry is proposed to improve the AFM measurement efficiency. In this method, the sample is measured firstly by white light scanning interferometry to get an overall coarse morphology, and then, further measured with higher resolution by AFM. Several measuring experiments on standard samples demonstrate the system’s good measurement performance and feasibility of the hybrid measurement method.

20 $\mu$A to 100 mA DC–DC Converter With 2.8-4.2 V Battery Supply for Portable Applications in 45 nm CMOS

A DC-DC buck converter capable of handling loads from 20 μA to 100 mA and operating off a 2.8-4.2 V battery is implemented in a 45 nm CMOS process. In order to handle high battery voltages in this deeply scaled technology, multiple transistors are stacked in the power train. Switched-Capacitor DC-DC converters are used for internal rail generation for stacking and supplies for control circuits. An I-C DAC pulse width modulator with sleep mode control is proposed which is both area and power-efficient as compared with previously published pulse width modulator schemes. Both pulse frequency modulation (PFM) and pulse width modulation (PWM) modes of control are employed for the wide load range. The converter achieves a peak efficiency of 75% at 20 μA, 87.4% at 12 mA in PFM, and 87.2% at 53 mA in PWM.

Local interaction imaging by SiGe quantum dot probe

Local interaction imaging of cleaved InAs surface (110) using a high aspect ratio (HAR) SiGe quantum dot (QD) probe was demonstrated by frequency-modulation mode noncontact atomic force microscopy under atmospheric pressure. The local contrast image with pronounced brightness is mainly attributed to no contact potential difference between the HAR SiGe-QD probe apex and the sample surface, and low frequency-change of the HAR SiGe-QD probe.

Electrostatic interaction forces in aqueous salt solutions of variable concentration andvalency

We use atomic force microscopy (AFM) to determine electrostatic interactions between Sitips and Si wafers in aqueous electrolytes of variable composition. We demonstrate thatdynamic force spectroscopy (DFS) in the frequency modulation (FM) mode with stiffcantilevers and sharp tips allows for a continuous detection of the tip–sample interactionswithout mechanical jump-to-contact instability and with substantially higher lateral resolutionthan standard colloidal probe measurements. For four different species of salt (NaCl, KCl,MgCl2,CaCl2) we find repulsive electrostatic forces at the lowest salt concentrations (1 mM) that becomeprogressively screened until they are dominated by attractive van der Waals forces atthe highest concentration (100 mM). For the divalent cations the crossover fromrepulsive to attractive forces occurs at lower concentrations than for monovalentcations. Surface charges extracted from fits to standard Poisson–Boltzmann doublelayer theory indicate a rather weak dependence of the surface charge on the ionconcentration. The high lateral resolution of our approach is illustrated by a 2D force fieldmeasurement over a patterned surface of a supported lipid bilayer on a mica substrate.

Metrological atomic force microscope with self-sensing measuring head

A metrological atomic force microscope (AFM) adapting compact measuring head for the micro-nano dimensional measurement is introduced in this paper. Based on a novel self-sensing probe the head does not need any optical detector to measure the bending of the cantilever, significantly saving the working space to integrate with other measuring methods. This AFM operates in tapping mode and adapts frequency modulation (FM) mode to enhance measuring speed. A z-axis piezoelectric positioning stage (compact z-stage), with high resonant frequency, is responsible for the rapid motion of the sample in z-direction. A high-end digital signal processing (DSP) servo control system further guarantees high measurement speed. A nano-measuring machine (NMM) is equipped as the lateral moving stage to realize three-dimensional measurement. Three interferometers in the NMM enable the measurement to be traced to the meter definition. After calibration, the measurement of one-dimensional grating has been made. According to experimental results, the resolution of the AFM can reach nanometer level.

J111023 非線形フィードバックによるFM-AFM用マイクロカンチレバープローブの振幅制御([J11102]マイクロナノ理工学:nmからmmまでの表面制御とその応用(2))

This study aims at improving the precision of measurements with a frequency modulation (FM) mode atomic force microscope (AFM). While an observation with the FM-AFM is carried out, a vibrating microcantilever probe scans over the sample surface. The measurement accuracy depends on how exactly a change in the eigen-frequency of the microcantilever is obtained. The amplitude suppression enables not only to make the oscillation frequency agree exactly with the eigen-frequency, but also to realize a non-contact observation for easily-deformable or easily-damageable specimens. However, it is difficult to cope with both keeping the microcantilever vibrating firmly and making the amplitude of its vibration as small as possible. To overcome this difficulty, a nonlinear feedback is applied to control the amplitude of the microcantilever probe used in the AFM. As a result, it is achieved that the AFM microcantilever can oscillate with a considerably reduced amplitude in the steady state. The key to the success of this nonlinear feedback control method is that the cantilever vibration can be categorized into the van der Pol-type self-excited oscillation. A prototype AFM was built to demonstrate experimentally the advantage of this amplitude control method. To date, the vibration amplitude less than one nanometer has been achieved in the steady state.

Implementation and characterization of a quartz tuning fork based probe consisted of discrete resonators for dynamic mode atomic force microscopy

The quartz tuning fork based probe {e.g., Akiyama et al. [Appl. Surf. Sci. 210, 18 (2003)]}, termed "A-Probe," is a self-sensing and self-actuating (exciting) probe for dynamic mode atomic force microscope (AFM) operation. It is an oscillatory force sensor consisting of the two discrete resonators. This paper presents the investigations on an improved A-Probe: its batch fabrication and assembly, mounting on an AFM head, electrical setup, characterization, and AFM imaging. The fundamental features of the A-Probe are electrically and optically characterized in "approach-withdraw" experiments. Further investigations include the frequency response of an A-Probe to small mechanical vibrations externally applied to the tip and the effective loading force yielding between the tip and the sample during the periodic contact. Imaging of an electronic chip, a compact disk stamper, carbon nanotubes, and Si beads is demonstrated with this probe at ambient conditions in the so-called frequency modulation mode. A special probe substrate, which can snap on a receptacle fixed on an AFM head, and a special holder including a preamplifier electronic are introduced. We hope that the implementation and characterization of the A-Probe described in this paper will provide hints for new scanning probe techniques.

Local dielectric spectroscopy of nanocomposite materials interfaces

Local dielectric spectroscopy is performed to study how relaxation dynamics of a polyvinyl-acetate ultrathin film is influenced by inorganic nanoinclusions of a layered silicate (montmorillonite). Dielectric-loss spectra are measured by electrostatic-force microscopy in the frequency-modulation mode in ambient air. Spectral changes in both shape and relaxation time are evidenced across the boundary between pure polymer and montmorillonite sheets. Dielectric-loss imaging is also performed, evidencing spatial variations of dielectric properties near nanostructures with nanometer-scale resolution.

Kelvin probe force microscopy in application to biomolecular films: Frequency modulation, amplitude modulation, and lift mode

Kelvin probe force microscopy (KPFM) is a powerful technique to visualize the differences of work function in metals and lateral surface potential distribution in thin organic films. Earlier we have shown that frequency modulated-Kelvin probe force microscopy has significant advantages in both sensitivity and resolution when applied to metal and inorganic interfaces in vacuum. High resolution, high sensitivity, and performance in ambient conditions are required in order to study biologically relevant samples. In this work we compared the resolution of frequency modulation (FM-KPFM), amplitude modulation (AM-KPFM), and lift modes KPFM for imaging the local electrical surface potential of complex biomolecular films and demonstrated that FM-KPFM mode has superior resolution for biological applications. The power of the method was illustrated on pulmonary surfactant films, revealing nm spatial resolution and mV potential sensitivity in ambient air. At this level of resolution this method can provide critical insight into the molecular arrangement and function of complex biosystems in a way that other KPFM modes cannot do. Based on the observed changes in the local surface potential we discovered that excess cholesterol produces nm size electrostatic domains and change the electric fields.

Effects of CRAC Channel on Spatiotemporal Ca2+ Patterns in T Cells

A reaction-diffusion model is built to investigate the temporal and spatial patterns of cytoplasmic Ca2+ dynamics under the effects of Ca2+-release activated Ca2+ (CRAC) channels in T cells. Simulation results show a strong dependence of the modulation mode of Ca2+ oscillation and dynamic patterns of Ca2+ wave on the influx rate through the CRAC channel (ksoc). When ksoc is small, cytoplasmic Ca2+ is modulated as a frequency-modulation (FM) signal, whereas it shows an amplitude modulation (AM) mode after ksoc passes through a critical value. The heterogeneity in spatial Ca2+ distribution is mostly arising from the influx through CRAC channels in both FM and AM modes. During each Ca2+ spike, a more sustained cytoplasmic Ca2+ gradient is maintained in the AM mode rather than in the FM mode.

Cytoplasmic Ca2+ Dynamics under the Interplay between the Different IP3R Gating Models and the Plasma Membrane Ca2+ Influx

The cytoplasmic Ca oscillations are investigated under the effect of CRAC channels in non-excitable cells (especially in T cells). The oscillatory Ca2+ signals can be modulated as the amplitude-, frequency- and mixed amplitude-frequency modulation modes dependent on the different IP3R gating models. Bifurcation analyses show that Ca2+ signals in the single positive feedback loop model is a mixed modulation mode. In contrast, Ca2+ signals in the Mak–McBride–Foskett model demonstrates approximately the frequency modulation mode only with slight amplitude shifts.

FM Signaling in Single Cells

Regulation of transcription factor localization allows cells to respond rapidly to extracellular signals. Although the molecular mechanisms of nuclear import and export have been examined, it remains unclear how localization varies among individual cells, and how dynamic changes in localization affect expression of downstream genes. In the presence of extracellular calcium, Crz1, the calcineurin responsive zinc finger transcription factor of Saccharomyces cerevisiae, is dephosphorylated and translocates into the nucleus. By observing the localization of Crz1-GFP fusion proteins using time-lapse microscopy, we found that Crz1 exhibited bursts of nuclear localization with a characteristic nuclear residence time of ∼2 minutes. These bursts occurred in a stochastic fashion in individual cells and propagated to the expression of downstream genes, contributing significantly to fluctuations in gene expression. Strikingly, calcium concentration controlled the frequency, but not duration, of nuclear localization bursts. Using an analytic model , we find that the observed stochastic frequency modulation (FM) of localization bursts can enable cells to proportionally coordinate expression levels of multiple target genes by regulating the fraction of time a promoter is active, rather than tuning the level of activity itself. We experimentally confirmed this theory by showing that both natural and synthetic Crz1 target promoters are expressed proportionally across a wide range of calcium concentrations. Furthermore, we observe that many proteins exhibit localization bursts and show diverse dynamic behaviors. These results suggest that cells may utilize FM mode of regulation to control diverse cellular processes.

In situ studies of the atomic layer deposition of thin HfO 2 dielectrics by ultra high vacuum atomic force microscope

We studied in situ the initial stages of atomic layer deposition (ALD) of HfO 2 by an ultra high vacuum atomic force microscope working in frequency-modulation mode. The ALD cycles, made by using tetrakis-di-methyl-amido-Hf and water as precursors, were performed on the Si(001)/SiO 2 substrate maintained at 230 °C. After each ALD cycle we studied the influence of the HfO 2 growth on the surface height histogram, the root mean square roughness, the surface fractal dimension and the autocorrelation function. This detailed analysis of the surface topography allowed us to confirm the completion of the first HfO 2 layer after four ALD cycles.

Spatiotemporal reconstruction of the auditory steady-state response to frequency modulation using magnetoencephalography

The aim of this study was to investigate the mechanisms involved in the perception of perceptually salient frequency modulation (FM) using auditory steady-state responses (ASSRs) measured with magnetoencephalography (MEG). Previous MEG studies using frequency-modulated amplitude modulation as stimuli (Luo et al., 2006, 2007) suggested that a phase modulation encoding mechanism exists for low (<5 Hz) FM modulation frequencies but additional amplitude modulation encoding is required for faster FM modulation frequencies. In this study single-cycle sinusoidal FM stimuli were used to generate the ASSR. The stimulus was either an unmodulated 1-kHz sinusoid or a 1-kHz sinusoid that was frequency-modulated with a repetition rate of 4, 8, or 12 Hz. The fast Fourier transform (FFT) of each MEG channel was calculated to obtain the phase and magnitude of the ASSR in sensor-space and multivariate Hotelling's T2 statistics were used to determine the statistical significance of ASSRs. MEG beamformer analyses were used to localise the ASSR sources. Virtual electrode analyses were used to reconstruct the time series at each source. FFTs of the virtual electrode time series were calculated to obtain the amplitude and phase characteristics of each source identified in the beamforming analyses. Multivariate Hotelling's T2 statistics were used to determine the statistical significance of these reconstructed ASSRs. The results suggest that the ability of auditory cortex to phase-lock to FM is dependent on the FM pulse rate and that the ASSR to FM is lateralised to the right hemisphere.

NMR stabilization of the magnetic field by a III1-1 magnetic inductometer in the frequency modulation mode

As applied to the problem of stabilizing the magnetic field of the NMR relaxometer-diffusiometer by a III1-1 magnetic inductometer, we propose a simple method for changing the III1-1 magnetic inductometer to the frequency modulation mode to exclude the field modulation arising from modulating coils of the III1-1 sensor at the studied sample location.

Response of Carbon Nanotube Field Effect Transistors to Vibrating Gate Determined by Scanning Gate Microscopy

The response of a carbon nanotube field effect transistor (CNT-FET) to a vibrating gate has been investigated by scanning gate microscopy (SGM) with a frequency modulation (FM) mode in air at room temperature. Topography and current modulation images were obtained simultaneously. The CNT-FET successfully detected a cantilever vibration with an amplitude less than 50 nm and afrequency of 80 kHz.