J. P. Cleveland

A nondestructive method for determining the spring constant of cantilevers for scanning force microscopy

The spring constant of microfabricated cantilevers used in scanning force microscopy (SFM) can be determined by measuring their resonant frequencies before and after adding small end masses. These masses adhere naturally and can be easily removed before using the cantilever for SFM, making the method nondestructive. The observed variability in spring constant—almost an order of magnitude for a single type of cantilever—necessitates calibration of individual cantilevers in work where precise knowledge of forces is required. Measurements also revealed that the spring constant scales with the cube of the unloaded resonant frequency, providing a simple way to estimate the spring constant for less precise work.

Tapping mode atomic force microscopy in liquids

Tapping mode atomic force microscopy in liquids gives a substantial improvement in imaging quality and stability over standard contact mode. In tapping mode the probe‐sample separation is modulated as the probe scans over the sample. This modulation causes the probe to tap on the surface only at the extreme of each modulation cycle and therefore minimizes frictional forces that are present when the probe is constantly in contact with the surface. This imaging mode increases resolution and reduces sample damage on soft samples. For our initial experiments we used a tapping frequency of 17 kHz to image deoxyribonucleic acid plasmids on mica in water. When we imaged the same sample region with the same cantilever, the plasmids appeared 18 nm wide in contact mode and 5 nm in tapping mode.

Measuring the viscoelastic properties of human platelets with the atomic force microscope

We have measured force curves as a function of the lateral position on top of human platelets with the atomic force microscope. These force curves show the indentation of the cell as the tip loads the sample. By analyzing these force curves we were able to determine the elastic modulus of the platelet with a lateral resolution of approximately 100 nm. The elastic moduli were in a range of 1-50 kPa measured in the frequency range of 1-50 Hz. Loading forces could be controlled with a resolution of 80 pN and indentations of the platelet could be determined with a resolution of 20 nm.

Short cantilevers for atomic force microscopy

We have designed and tested a family of silicon nitride cantilevers ranging in length from 23 to 203 μm. For each, we measured the frequency spectrum of thermal motion in air and water. Spring constants derived from thermal motion data agreed fairly well with the added mass method; these and the resonant frequencies showed the expected increase with decreasing cantilever length. The effective cantilever density (calculated from the resonant frequencies) was 5.0 g/cm3, substantially affected by the mass of the reflective gold coating. In water, resonant frequencies were 2 to 5 times lower and damping was 9 to 24 times higher than in air. Thermal motion at the resonant frequency, a measure of noise in tapping mode atomic force microscopy, decreased about two orders of magnitude from the longest to the shortest cantilever. The advantages of the high resonant frequency and low noise of a short (30 μm) cantilever were demonstrated in tapping mode imaging of a protein sample in buffer. Low‐noise images were tak...

Mapping interaction forces with the atomic force microscope

Force curves were recorded as the sample was raster-scanned under the tip. This opens new opportunities for imaging with the atomic force microscope: several characteristics of the samples can be measured simultaneously, for example, topography, adhesion forces, elasticity, van der Waals, and electrostatic interactions. The new opportunities are illustrated by images of several characteristics of thin metal films, aggregates of lysozyme, and single molecules of DNA.

Studies of vibrating atomic force microscope cantilevers in liquid

An atomic force microscope (AFM) design providing a focused spot of order 7 μm in diameter was used to analyze the motion of vibrating cantilevers in liquid. Picking an operating frequency for tapping mode AFM operation in liquid is complex because there is typically a large number of sharp peaks in the response spectrum of cantilever slope amplitude versus drive frequency. The response spectrum was found to be a product of the cantilever’s broad thermal noise spectrum and an underlying fluid drive spectrum containing the sharp peaks. The geometrical shape of transverse cantilever motion was qualitatively independent of the fluid drive spectrum and could be approximately reproduced by a simple theoretical model. The measurements performed give new insights into the behavior of cantilevers during tapping mode AFM operation in liquid.

PROTEIN TRACKING AND DETECTION OF PROTEIN MOTION USING ATOMIC FORCE MICROSCOPY

Height fluctuations over three different proteins, immunoglobulin G, urease, and microtubules, have been measured using an atomic force microscope (AFM) operating in fluid tapping mode. This was achieved by using a protein-tracking system, where the AFM tip was periodically repositioned above a single protein molecule (or structure) as thermal drifting occurred. Height (z-piezo signal) data were taken in 1 - or 2-s time slices with the tip over the molecule and compared to data taken on the support. The measured fluctuations were consistently higher when the tip was positioned over the protein, as opposed to the support the protein was adsorbed on. Similar measurements over patches of an amphiphile, where the noise was identical to that on the support, suggest that the noise increase is due to some intrinsic property of proteins and is not a result of different tip-sample interactions over soft samples. The orientation of the adsorbed proteins in these preliminary studies was not known; thus it was not possible to make correlations between the observed motion and specific protein structure or protein function beyond noting that the observed height fluctuations were greater for an antibody (anti-bovine IgG) and an enzyme (urease) than for microtubules.

Noncontact force microscopy in liquids

Force microscopy in liquids offers many advantages including the mitigation of capillary forces and the simulation of real environments for biological and technological processes. Noncontact force microscopy in liquids adds the advantage of probing electrical and magnetic fields above surfaces. Here we demonstrate magnetic force imaging of recorded bits on a computer hard disk in air and in liquid. A method of noncontact force microscopy (patent pending, Digital Instruments) is used in which the tip is first scanned in contact to image topography and then rescanned above the surface to image long‐range forces.Force microscopy in liquids offers many advantages including the mitigation of capillary forces and the simulation of real environments for biological and technological processes. Noncontact force microscopy in liquids adds the advantage of probing electrical and magnetic fields above surfaces. Here we demonstrate magnetic force imaging of recorded bits on a computer hard disk in air and in liquid. A method of noncontact force microscopy (patent pending, Digital Instruments) is used in which the tip is first scanned in contact to image topography and then rescanned above the surface to image long‐range forces.

Lattice resolution and solution kinetics on surfaces of amino acid crystals: an atomic force microscope study

Abstract We report atomic force microscopy (AFM) results on six amino acid crystal surfaces: glycine, L-aspartic acid, L-valine, L-isoleucine, L-leucine, and L-phenylalanine. Samples were grown by slow evaporation of concentrated aqueous solutions. All samples contained crystalline areas where the AFM showed extended molecularly flat sheets (up to hundreds of nm in size) separated by steps a single molecule thick. The ordered lattice of each amino acid could be imaged on the sheets. Images revealed periodicities corresponding to bulk terminations in most cases, as well as other periodicities which probably correspond to molecular structure within the unit cell. Step motion kinetics were also imaged in situ during dissolution of L-leucine in flowing propanol. Steps oriented along the 〈010〉 direction traveled with speeds that were independent of both interstep distance and solvent flow rate for flow rates above 20 μl/s, indicating a reaction rate limited process. Orthogonal bends along the 〈001〉 direction moved at speeds one to ten times that of steps, with narrow bends moving faster than wide. We speculate that these speed differences were caused by anisotropy in reaction kinetics coupled with partially saturated boundary layers near wide bends.

Atomic force microscope for small cantilevers

We have designed and built an atomic force microscope (AFM) with optical beam deflection detection providing a focused spot size of 1.6 micrometers in diameter. This small spot size was implemented with a variable focus adjustment that allows us to re-focus on each cantilever. This design opens up the usage of a new range of small cantilevers with low-noise characteristics. We have microfabricated novel aluminum cantilevers with dimensions as small as 9 micrometers in length and 2.5 micrometers in width and have characterized them with this new AFM. The resonance frequency of the smallest cantilever was 2.5 MHz in air and 0.94 MHz in water. We demonstrated the imaging capabilities of the AFM head by imaging abalone nacre with a 10 micrometers long cantilever using the tapping mode in liquid at a drive frequency of 442 KHz.