Atomic force microscopy,

The atomic force microscope can be configured in several ways, the most obvious (contact mode) merely involving scanning the tip over the sample at regular intervals, rasper fashion, and recording the deflection. Because of the proportionately very large capillary forces that arise from a contamination layer, imaging of polysaccharides in direct contact mode is carried out under a solvent 

The ability of the tip to pick up one end of a biopolymer while the other still remains bound to the base has permitted direct measurements of forces 

Modification of the atomic force microscope with feedback loops enabled force-extension curves for individual polysaccharide molecules to be measured under constantly increasing force (rather than constant displacement), in a nanoscale version of the INSTRON tester for measuring paper strength.  

With rare exceptions (one discussed above), the monosaccharide units, particularly pyranose units, of oligosaccharides and polysaccharides can be regarded as rigid. The conformation of any inter-saccharide linkage can then be 

It is widely observed that preferred conformations of glycosides with equatorial glycosidic C-O bonds have values of (pn in the general range 45-25° ((p=-15 to -95°) and those of axial glycosides around -30° (cp = 90°). The electronic or electrostatic interactions that determine these preferences are the same as those governing the anomeric effect and were termed the exo-anomeric effect by Lemieux and co-workers.Whereas the no-bond resonance explanation for 

Atomic force microscopy (AFM) allows the topography of a sample to be scanned by using a very small tip made from silicon nitride. The tip is attached to a cantilever that is characterised by its spring constant, resonance frequency, and a quality factor. The sample rests on a piezoceramic tube which can be moved horizontally x,y motion) and vertically (z motion). Displacement of the cantilever is measured by the position of a laser beam reflected from the mirrored surface on the top side of the cantilever, whereby the reflected laser beam is detected by a photodetector. AFM can be operated in either contact or a noncontact mode. In contact mode the tip travels in close contact with the surface, whereas in noncontact mode the tip hovers 5-10 nm above the surface. 

Atomic force microscopy (AFM) has become a standard technique to image with high resolution the topography of surfaces. It enables one to see nanoscopic surface features while the electrode is under potential control. This powerful probe microscopy operates by measuring the force between the probe and the samples (56,57). The probe consists of a sharp tip (made of silicon or silicon nitride) attached to a force-sensitive cantilever. The tip scans across the surface (by a piezoelectric scanner), and the cantilever deflects in response to force interactions between the tip and the substrate. Such deflection is monitored by bouncing a laser beam off it onto a photodetector. The measured force is attributed to repulsion generated by the overlap of the electron cloud at the probe tip with the electron cloud of surface atoms. 

Atomic force microscopy (AFM) has been used to characterize dendrimers that have been adsorbed onto a surface such as silica. AFM involves moving a finely tipped stylus across a surface and monitoring the tip movements as it traces the surface topography. In studying adsorbed dendrimers, samples can be scanned repeatedly and in a variety of directions. When this is done, it is found that all the images are the same. True dendrimers form objects of only one size. 

AFM has also been used to study adsorbed dendrimers of increasing generation number. When this is done, it is found that there is a steady and regular rise in the diameters of the adsorbed dendrimers with each additional generation number, but that each generation of dendrimers remains highly homogeneous. 

Atomic force microscopy (AFM) has become a standard technique for high-resolution imaging of the topography of surfaces. It enables one to see nanoscopic 

FIGURE 2-15 Design of a system for in-situ electrochemical scanning tunneling microscopy. 

Atomic force microscopy (AFM) is a microscopic method for chemists and biologists offering the magnification range of both the light and electron 

The concept of resolution in AFM is different from radiation-based microscopies because AFM is a three-dimensional imaging technique. There is an important distinction between images resolved by wave optics and scanning probe techniques. The former is limited by diffraction, whereas the latter is limited primarily by apical probe geometry and sample geometry. Usually the width of a DNA molecule is loosely used as a measure of resolution, because it has a known diameter of 2.0 nm in its B form. 

The first highly reproducible AFM images of DNA were obtained in 1991. Four major advances that have enabled clear resolution of nucleic acids are 

Atomic force microscopy has been developed to a stage that DSBs and the length of the resulting fragments can be detected by this technique (Pang et al. 1996). In this context it is of interest that neutron irradiation leads to the formation of very small fragments (Pang et al. 1997). 

Atomic force microscopy has been used to evaluate the nature and size of the polyurethane structure at the surface of a sample. In this method, the surface of the sample is coated with a layer of gold and a probe scans over the surface. As the nature of the chemistry below the probe changes, the force field acting on the probe changes. The hard segment clusters can be visualized. 

Atomic force microscopy has been combined with nano-indentation measurements to map hardness variations on the surface of a CaCOg-filled sample of PDMS. ° In another application, PDMS-modified tips were used to obtain friction coefficients involving self-assembled monolayers.  

Atomic force microscopy is a rather new new method to characterise the surface of a membrane [6,7]. A sharp tip with a diameter smaller than 100 A is scanning across a surface with a constant force. London - vanderWaals interactions will occur between the atoms in the tip and the surface of the sample and these forces are detected. This will result 

Atomic force microscopy (AFM) allows material characterization (conductive or not) down to the nanoscale, in environmental conditions [20, 21]. It was developed 

Atomic force microscopy (AFM) maps out the topographical features of a surface. A miniature tip (radius of curvature is 5-10 nm °), usually made of silicon or silicon nitride, taps along a prescribed area of the surface being imaged.Attractive and repulsive electrostatic forces affect the motion of the tip as it operates close to the surface. A piezoelectric scanning actuator works in a feedback loop to raise or lower the 

Schematic of an atomic force microscope (a) and the force profile being sensed during the scan (b). 

Whilst the above methods can provide a visualization of the surface topography, they are unable to address two questions. Firstly, they are unable to provide information on the density distribution relative to the notional surface, and secondly, they are unable to allow identification of the surface elemental, atomic, composition. 

4 Spectroscopic Assessment of the Surface Attenuated Total Reflection Infrared, Fluorescence and Visible Spectroscopy  

In the context of morphology, the principal information that is obtained from such experiments is the conformational distribution of the polymer chains. In recent years, microscopes have been developed which allow the simultaneous observation of the surface and spectroscopic examination of the surface. The illuminating beam is brought down the optical axis and the reflected light is then collected using fibre optics. The spot size is typically several tens of micrometres but can be smaller and does allow characterization of domains or phase structure that has dimensions of this order. The techniques and their application are covered in detail elsewhere.  

7 OTHER INSTRUMENTAL METHODS 3.7.1 Atomic Force Microscopy 

One of the most popular choices of microscopy is atomic force microscopy (AFM), which is a modification of the earlier scanning tunnelling microscope 

Principles and Characteristics In 1986 Binnig et al. [292] have developed the atomic force microscope (AFM) which remedied a severe limitation of STM, namely imaging of conducting materials only. The first commercially available AFM was introduced in 1989. Since that time, AFM has been used with great success to study surfaces of insulators and the macromolecular architecture of polymeric materials from sub-nm to ptm scale [285,286,293]. 

AFM is essentially a very sensitive profilome-ter. In AFM an atomically sharp stylus or probe (a few /u,m long and often less than 5 nm in diameter. 

Since its inception by Binnig et al. in 1986, AFM has become an important and widespread tool for 

This procedure keeps the tip in the region where the tip-sample force is (relatively) well understood, but at the price that the force is determined by the cumulative effect of a large number of atoms - hence the resolution of individual atomic-scale features is seldom possible. In the non-contact mode, the cantilever is made to vibrate at its resonant frequency, and the interaction damps the amplitude of the vibration. NC-AFM is preferable to contact AFM for measuring soft samples such as polymers, but the spatial resolution is lower. 

Because the interactions between tip and surface depend not only on the topography of the sample but also on different characteristics (such as hardness. 

In a recent study Raj et al. presented the first direct study of adhesion forces, by colloidal force microscopy, between smooth PLA films representing the polymer matrix, and a microbead of cellulose that mimics the cellulose material in flax fibers [65]. Normalized adhesion force measurements demonstrated the importance of capillary forces when experiments were carried out under ambient conditions. Experiments, conducted under dry air allowed for the deduction of the contribution of pure van der Waals forces, and the results, through the calculation of the Hamaker constant, show that these forces, for the PLA/cellulose/air system, were lower than those obtained for the cellulose/cellulose/air system and hence underlined the importance of optimizing the interface among these materials. The study demonstrated the capacity of AFM to probe direct interactions in complex systems by adjusting the nature of the surface and 

Contact mode Being the most basic mode of operation, contact mode is widely used. The tip is raster-scanned across the surface, where it is deflected following surface corrugation. In constant-force mode, the tip is constantly adjusted to maintain a fixed deflection, thus keeping a constant height above the surface this adjustment along the z axis is displayed as data. However, the ability to track the surface in this way is limited by the response of the 

Electron-matter interaction used for analytical microscopy 

Transmitted electrons with elastic dispersion (energy ) 

Transmitted electrons with inelastic dispersion energy -A EELS) 

Repulsive force tip is pushed away I from the surface 

Microthermal analysis (MTA) is a technique that combines thermal analysis and atomic force microscopy (AFM). This technique has been discussed in a series of papers published by Slough and co-workers in 1999 [1 ] and by other researchers [2-7]. 

MTA is now being used commercially to visualise the spatial distribution of phases, components and contaminants in polymers, foods biological materials and electronic materials. Pollock and Hammiche [4] review various applications that have been described in the literature to date, ranging from multi-layer materials and interphase regions in composites, to the use of the regions technique as a means of surface treatment in this paper. 

The force microscope, in general, has several modes of operation. In the repulsive-force or contact mode, the force is of the order of 1-10 eV/A, or 10 -10 newton, and individual atoms can be imaged. In the attractive-force or noncontact mode, the van der Waals force, the exchange force, the electrostatic force, or magnetic force is detected. The latter does not provide atomic resolution, but important information about the surface is obtained. Those modes comprise different fields in force microscopy, such as electric force microscopy and magnetic force microscopy (Sarid, 1991). Owing to the limited space, we will concentrate on atomic force microscopy, which is STM s next of kin. 

The AFM has a number of elements common to STM the piezoelectrc scanner for actuating the raster scan and z positioning, the feedback electronics, vibration isolation system, coarse positioning mechanism, and the computer control system. The major difference is that the tunneling tip is replaced by a mechanical tip, and the detection of the minute tunneling current is replaced by the detection of the minute deflection of the cantilever. 

In order to achieve sufficient sensitivity for atomic resolution, the cantilever has to satisfy several requirements (Albrecht et al., 1990). 

the cantilever must be flexible yet resilient, with a force constant from 10 to 10 N/m. Therefore, a change of force of a small fraction of a nanonewton (nN) can be detected. 

Second, the resonance frequency of the cantilever must be high enough to follow the contour of the surface. In a typical application, the frequency of the corrugation signal during a scan is up to a few kHz. Therefore, the natural frequency of the cantilever must be greater than 10 kHz. 

In general, the STM s ability to obtain atom images is clearly outstanding. Unfortunately, there are some limitations, particularly the requirements that both the tip and sample surface are good conductors. This severely limits the materials that can be studied and has led to the development of scanning force microscopy (atomic force microscopy) which is described in Section 5.3. 

Many of the observations have been made on replicas of rotary shadowed structures that had been dried on the carrier. These dehydrated samples have most obviously lost some of their physiological properties beside the fact that the preparation as such could be deleterious to the sample and result in artefacts. The development of atomic force microscopy (AFM) in the 1980s is therefore a valuable improvement of technology towards more physiological sample application possibilities. 

In this section we summarize experimental methods that enable measuring (depletion) interaction potentials between particles [64]. We distinguish pair interactions (Sects. 2.6.1-2.6.3) and many-body interactions (Sect. 2.6.4). The latter can be measured indirectly using scattering techniques or microscopy, whereas for pair interactions direct methods are available. Common instruments for investigating such pair interactions are the surface force apparams (SFA) [65], optical tweezers [66, 67], atomic force microscopy (AFM) [68], and total internal reflection microscopy (TIRM) [69, 70]. 

The effective pair interactions measured with these techniques are the direct pair interactions between two colloidal particles plus the interactions mediated by the depletants. In practice depletants are poly disperse, for which there are sometimes theoretical results available. For the interaction potential between hard spheres we quote references for the depletion interaction in the presence of polydisperse penetrable hard spheres [74], poly disperse ideal chains [75], poly-disperse hard spheres [76] and polydisperse thin rods [77]. 

The atomic force microscope (AFM) was designed for high-resolution surface topography analysis. The basic measuring principle is sketched in Fig. 2.34. A sample is scanned by a sharp tip attached to a sensitive cantilever spring via a 

Interactions between a spherical colloid and a wall can be measured by bringing probe and substrate together and monitoring the cantilever deflection as a function of the interparticle distance. The photodetector voltage versus piezo position curve can be converted into a force-distance curve. The force acting on the cantilever follows from the deflection of the cantilever and its known spring constant. The zero force is defined by the deflection of the cantilever as the colloidal probe is far from the surface of the substrate. To obtain the force-distance dependence on an absolute scale the zero distance, i.e., where the colloid touches the wall, has to be determined. Commonly, the zero distance is obtained from the force curve itself and not through an independent method [68]. 

In practice, the position where the motion of the probe complies with the piezo movement defines the point of zero distance. Force-distance curves recorded with AFM depend on the specific geometry of the probe and the surface. Usually, the interaction is displayed as the force divided by the radius of the colloid, R, in units N/m. The Derjaguin approximation relates this quantity to the interaction potential per unit area between equivalent flat surfaces at given separation distance, see (2.29). 

AFM is an advanced tool that is ideal for examination of microscopic surface topography. The main advantages of AFM over profilometry are its 

AFM can be run in three different modes contact, noncontact, and tapping mode. When AFM is in the contact mode (similar to stylus profilometry), the most common problem encountered is that under ambient conditions, sample surfaces are covered by a layer of adsorbed gases consisting primarily of water vapor and nitrogen. In addition, a dielectric film can trap electrostatic charge, which can contribute to additional attractive forces between probe and sample. These problems may cause friction in probing, which will destroy the sample or distort the resulting data. 

Differences between noncontact and tapping mode AFM, The signal for the former is dependent on the change in oscillation due to the force gradient, while the latter is dependent on the oscillation change due to the contact, (Courtesy of Digital Instruments, Veeco 

AFM Surface Roughness Comparison (RMS Stands for Root Mean Square) 

Position Slurry A (nm) Slurry B (nm) Slurry C with buff (nm) 

AFM has been used to detect surface topological changes on the imprinted compounds that occur when the imprinted componnds bind the analytes. However, AFM is very slow and the method is impractical nnless the chip can be scanned with a large array of AFM probes in parallel. 

Like STM, AFM produces 3-D images. Unlike STM, AFM does not require the sample to be conducting and so has wider applicability. Unlike optical microscopes, AFM does not use a lens, so the resolution of the technique is limited by probe size (and strength of interaction between surface and probe tip) rather than by diffraction effects. The key to the sensitivity of AFM is in carefully monitoring the movement of the probe tip. 

As in all SPM techniques, sample preparation is key. An AFM can work either when the probe is in contact with a surface, or when it is a few nanometres away. AFM can also be carried out on liquids. 

The discriminating element in the AFM is the cantilever and tip. When the tip moves over the surface, the cantilever moves and so too does the laser. The negative feedback loop moves the sample up and down via a piezoelectric scanner so as to maintain the interactive force. Typical AFM probe tips are made of silicon or silicon nitride, which provide lateral resolutions of 5-10 nm and 10-40 nm, respectively. There are three main modes of AFM and all depend on this interaction between the tip and the sample  

Any deflection in the laser is picked up by the photodiode detector and is converted into topographical information about the sample. 

The detector sends a signal to the computer for analysis and image generation. Information Obtained 

For simple imaging applications the AFM tip may be dragged across the surface as described above (the so called contact or static mode) or, alternatively, vibrated up and down near its resonant frequency in the z-direction ( acoustic or tapping mode). The tapping mode gives additional information about the elastic and other mechanical properties of the material being scanned. 

With suitably sharp AFM tips, almost atomic resolution can be achieved, at least in principle, and it has been used to determine the shapes of protein and nucleic acid molecules adsorbed on surfaces. One major advantage is that the method does work under water, so that molecules can be studied under near physiological conditions. 

Q In a typical AFM experiment to mechanically unfold a single globular protein, the cantilever arm moved by 25 nra with an average force of 150 pN as the polypeptide chain unravelled. How much mechanical work was done in the process How does this compare to the Gibbs free energy of unfolding  

Under ideal, thermodynamically reversible conditions, this mechanical work is equal to the Gibbs free energy change (AG) for the process. The numbers obtained here cannot be compared directly with the values obtained for molecules in solution using bulk thermodynamic methods (Chapter 5) because they include the additional elastic work involved in stretching the unfolded polypeptide as the tethered ends are pulled further apart than they would normally be for an unfolded protein free in solution. 

Scanning tiiniiclling microscopy (STM) works in a rather similar way to AFM, but the surface is detected by measuring the very weak electric current that flows (by electron tunnelling) between the sample and the tip when they are very close together with a voltage between them. Unlike AFM, this cannot be done under water. 

The complexity of the solid state offers not only an analytical challenge but also interesting aspects and opportunities for the dmg development. Thus, a drug can be engineered at the supramolecular and particulate level in order to optimize its physical properties. This requires, of course, a substantial knowledge of its solid-state properties and the proper analytical tools. 

(1988) Pharmaceutical Preformulation The Physicochemical Properties of Drug Substances, Ellis Horwood, Chichester. 

Lipinski, C.A., Lombarda, F., Dominy, B.W. Feeney, P.J. (2001) Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings, Adv. Drug Deliv. Rev., 46, 3-26. 

(1992) Crystal engineering and particle design for the powder compaction process. Drug Dev. Ind. Pharm., 18, 677-721. 

Padden, B.E., Munson, EJ. Grant, DJ.W. (1998) Solid-state characterization of two polymorphs of aspartame hemihydrate, J. Pharm. Set, 87, 501-507. 

The past decade has witnessed an explosion of techniques used to pattern polymers on the nano- and sub-micrometre scale, driven by the extensive versatility of polymers for diverse applications such as molecular electronics, data storage, and all forms of sensors. Lyuksyutov and co-workers [349] demonstrate a novel lithography technique - electrostatic nanolithography using AFM - that generates features by mass transport of polymer within an initially uniform, planar film without chemical crosslinking, substantial polymer degradation or ablation. 

The application of AFM and other techniques has been discussed in general terms by several workers [350-353]. Other complementary techniques covered in these papers include FT-IR spectroscopy, Raman spectroscopy, NMR spectroscopy, surface analysis by spectroscopy, GC-MS, scanning tunnelling microscopy, electron crystallography, X-ray studies using synchrotron radiation, neutron scattering techniques, mixed crystal infrared spectroscopy, SIMS, and XPS. Applications of atomic force spectroscopy to the characterisation of the following polymers have been reported polythiophene [354], nitrile rubbers [355], perfluoro copolymers of cyclic polyisocyanurates of hexamethylene diisocyanate and isophorone diisocyanate [356], perfluorosulfonate [357], vinyl polymers 

The application of atomic force microscopy to surface morphological studies has been covered in terms of the following polymers polyesters, PE, PS [369], polycarbonate, polyimide, PTFE 

AFM has been used in studies of adhesion in the cases of PS/PMMA [383], PET/PP [384], and polyurethane- and epoxy-based adhesives [385]. General discussions of the use of this technique in adhesion studies have been reported [355, 386]. 

Zhang and co-workers [387] in their discussion of the polydispersivity of ethylene sequence length in metallocene ethylene-a-olefin copolymers characterised by the thermal fractionation technique used AFM to study crystal morphology. 

The sample is mounted on a piezoceramic scanner with x,y,z movements small deflections of the cantilever are determined with an optical beam deflection system 

FIGURE 1.32 (a) Schematic representation of the experimental setup for SECM-AFM 

The AFM can be operated in several modes that are sufficiently different that they need to be described separately. The basic distinction between these modes is how much the tip touches the specimen surface contact mode is when the tip is in contact with the specimen all the time intermittent contact mode is when the tip oscillates and touches the specimen some of the time in noncontact mode, the tip does not touch the specimen at all. 

The lateral forces result in a torsion of the cantilever, and this can be measured using the same optical detection system that measures vertical deflection (see Fig. 2.8). This signal from the tip-specimen interaction leads to another operational mode, the lateral or frictional force microscope (LFM or FFM) [142, 143]. In LFM the cantilever is designed to be 

The concept of measuring forces was further developed in an operational mode called force spectroscopy. Scanning is typically discontinued and the experiment is performed at a given x, y location on the surface. Again, the cantilever support is moved vertically but over a much wider range. Measuring the cantilever deflection gives a force versus distance curve, which is a plot of the force (or cantilever deflection) as a function of the tip-sample separation. In particular, this method can be used to measure the pull-off force (i.e. the force required to separate the tip from the sample surface). In 

Another mode of operation derived from contact AFM that is relevant to polymer studies is the scanning thermal microscope (SThM) [118, 146], In SThM the tip is a special device that has a resistive element. If a current is passed through this resistive element, its temperature depends on the heat transferred to the specimen. It thus acts as a thermal probe, as seen for example in Fig. 5.36. During scanning, thermal control of the probe can be used to generate images based on variation of either sample temperature or thermal conductivity. Filled polymers and polymer blends are candidates for this kind of study, but the resolution is relatively poor. Microfabricated thermal probes can give a resolution of 100 nm. 

The application of contact mode AFM to soft materials such as polymers and biological systems is seriously limited because of the 

FIGURE 14.13 Schematic of simultaneous AFM-SECM imaging and the reactions involved at the surface of the micropattemed sample with integrated electrode operation in GC mode. (Reprinted with permission from Ref. [45]. Copyright 2003, Wiley-VCH Verlag GmbH.) 

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The ability to control the position of a fine tip in order to scan surfaces with subatomic resolution has brought scanning probe microscopies to the forefront in surface imaging techniques. We discuss the two primary techniques, scanning tunneling microscopy (STM) and atomic force microscopy (AFM) the interested reader is referred to comprehensive reviews [9, 17, 18]. 

AFM Atomic force microscopy [9, 47, 99] Force measured by cantilever deflection as probe scans the surface Surface structure... 

Friction can now be probed at the atomic scale by means of atomic force microscopy (AFM) (see Section VIII-2) and the surface forces apparatus (see Section VI-4) these approaches are leading to new interpretations of friction [1,1 a,lb]. The subject of friction and its related aspects are known as tribology, the study of surfaces in relative motion, from the Greek root tribos meaning mbbing. 

We have considered briefly the important macroscopic description of a solid adsorbent, namely, its speciflc surface area, its possible fractal nature, and if porous, its pore size distribution. In addition, it is important to know as much as possible about the microscopic structure of the surface, and contemporary surface spectroscopic and diffraction techniques, discussed in Chapter VIII, provide a good deal of such information (see also Refs. 55 and 56 for short general reviews, and the monograph by Somoijai [57]). Scanning tunneling microscopy (STM) and atomic force microscopy (AFT) are now widely used to obtain the structure of surfaces and of adsorbed layers on a molecular scale (see Chapter VIII, Section XVIII-2B, and Ref. 58). On a less informative and more statistical basis are site energy distributions (Section XVII-14) there is also the somewhat laige-scale type of structure due to surface imperfections and dislocations (Section VII-4D and Fig. XVIII-14). 

We confine ourselves here to scanning probe microscopies (see Section VIII-2B) scanning tunneling microscopy (STM) and atomic force microscopy (AFM), in which successive profiles of a surface (see Fig. VIII-1) are combined to provide a contour map of a surface. It is conventional to display a map in terms of dark to light areas, in order of increasing height above the surface ordinary contour maps would be confusing to the eye. 

The most popular of the scanning probe tecimiques are STM and atomic force microscopy (AFM). STM and AFM provide images of the outemiost layer of a surface with atomic resolution. STM measures the spatial distribution of the surface electronic density by monitoring the tiumelling of electrons either from the sample to the tip or from the tip to the sample. This provides a map of the density of filled or empty electronic states, respectively. The variations in surface electron density are generally correlated with the atomic positions. 

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