The Tools Of Nanomaterials Biology Essay

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The world of nanoscale materials is a constant bustle of activity. Nanoscale objects absorb, emit, make and break bond, vibrate and travel; they are always active. It is difficult to catch them and also nano-size objects are too small to observe with naked eye. Studying these tiny structures require special and deviously clever instruments that measure some properties of matter. Detecting and analyzing the light absorbed or emitted by atoms or molecules is the science of spectroscopy like Infrared, Raman and UltraViolet-Visible spectroscopy. Energy level transitions, energy and wavelength of electromagnetic spectrum are depicted in the electromagnetic ruler (Figure 5.1).

In the mid-1980's a number of high resolution microscopic instruments have been invented, which help the advancement of nanotechnology to a great extend. These microscopic techniques include scanning probe and electron microscopic techniques. The scanning tunneling microscope and atomic force microscope are examples of scanning probe microscopic technique whereas transmission electron microscope and scanning electron microscope are examples of electron microscopic technique. The discovery of these instruments has lead to numerous inventions on fundamental science. These high resolution instruments have also improved the synthesis of nanostructures by bottom up approach, where the structures are made by assembling atoms and molecules. The intention of this chapter is to provide the basic information about the fundamentals of various structural characterization tools, such as various spectroscopic and microscopic techniques, that are most widely used in characterizing nanomaterials and nanostructures.

5.2 X-ray Diffraction (XRD)

XRD is a very good tool for the determination of crystallinity, crystal structures and lattice constants of nanoparticles, nanowires and thin films. This experimental technique is also being used for applications, such as materials identification. XRD is the best method to examine whether a resultant material is amorphous or crystalline in nature. Crystalline phases can be identified by comparing the 'd' values obtained from XRD data with the fundamental data in Joint Committee on Powder Diffraction Standards (JCPDS).

A narrow beam of X-rays from a source is made incident on a specimen at an angle θ with the crystal plane. The wavelength of X-rays is usually ranging from 0.7 to 2 Ǻ. The beam is diffracted by the crystalline phases of the specimen. By Bragg's law:

2d sinθ = nλ, (n=1, 2, 3,…) (5.1)

where d is the interplanar spacing and λ is the wavelength of the incident X-rays. The intensity of the diffracted beam depends on the diffraction angle 2θ and the specimen's orientation. The study of diffraction pattern helps to measure specimen's structural properties because each sample produces X-rays of definite wavelengths which are characteristic of that material.

XRD is a suitable tool for characterizing homogeneous and inhomogeneous strains. From XRD pattern, one can easily measure diffraction peak positions. The diffraction peak positions will be shifted due to homogeneous or uniform elastic strain. From this shift, the change in interplanar spacing d can be calculated. The variation of d-spacing is due to the variation of lattice constants under a strain. The diffraction peaks are usually broadened and this broadening are mainly due to innhomogeneous strains. The innhomogeneous strains change from crystals to crystals or within a crystal and this is the cause of peak broadening. The broadening increases with sinθ. Another reason for peak broadening is the finite size of crystals and in such cases, the broadening is independent of sinθ. Careful analysis of peak shapes is necessary to find the contribution of both inhomogeneous strain and crystallite size to the peak width.

The size D of the crystallite can be calculated from the width of peak using Scherrer's formula, provided there is no inhomogeneous strain.

D =Kλ/(B cosθB), (5.2)

where K is Scherrer's constant, λ is the wavelength of X-rays, B is the full width at half maximum (FWHM) of the diffraction peak and θB is the diffraction angle. XRD of low Z elements are less sensitive as compared to high Z elements because the intensity of X-rays diffracted from low Z elements is low. XRD commonly requires large specimens because of small diffraction intensities. The information acquired in XRD is an average over a large quantity of materials. This technique is very useful for nanoparticle characterizion because it works only at very small dimensions. XRD can also be used to estimate the thickness of thin films.

5.3 Infrared (IR) Spectroscopy

IR spectroscopy is an important tool for sample identification and structural elucidation. This spectroscopic technique measures absorption of various IR frequencies by a specimen. IR spectroscopy with suitable sampling accessories helps us to use solid, liquid and gas samples.

Each atom in a molecule vibrates as the temperature is above absolute zero. When the frequency of vibration and the frequency of the incident IR beam are equal, the molecule absorbs the radiation. The method of analyzing this vibration is called infrared spectroscopy. Each atom has 3 degrees of freedom, corresponds to motion along any of the three coordinate axes (x, y, z). If a polyatomic molecule consists of n atoms, each atom has 3n degrees of freedom. Three of the degrees of freedom are used to describe translatory motion and another 3 degrees of freedom are used to describe the rotation of the molecule. The remaining (3n - 6) degrees of freedom are fundamental vibrations for nonlinear molecules. However, linear molecules have (3n - 5) fundamental modes of vibration since they require only 2 degrees of freedom to describe rotation. Only those fundamental modes of vibration, that produce a net change in the dipole moment can generate IR activity. The fundamental modes which cause polarizability changes can result Raman activity. Obviously, some vibrations are both IR and Raman active.

Fig. 5.1: Electromagnetic radiation ruler.

When IR radiation is incident on a sample, some frequencies are absorbed by the sample and some other frequencies are transmitted. The resulting spectrum consists of absorption peaks, corresponding to the frequencies of vibrations between the bonds of the atoms. It is significant that IR spectrum of different materials is different since each material is a unique combination of atoms. Clearly, IR spectroscopy gives a qualitative analysis of materials. Apart from this, IR spectroscopic technique gives a measure of the amount of materials present in the sample because the size of the absorption peaks is a direct indication of quantitative analysis of materials. The modern software based IR spectroscopy work as an excellent tool for quantitative analysis of materials. Figure 5.2 shows the sketch of a typical IR spectrometer.

Fig. 5.2: Schematic representation of IR Spectroscope

Rradiation source, monochromator and detector are the main components of IR spectrometer. Generally, an electrically heated inert solid (1000-1800 °C) is used as the radiation source. The monochromator disperse the wide spectrum of radiation and offers IR radiation of suitable frequency range. For this, a combination of prisms or gratings with variable slit mechanisms, mirrors and filters is used. Photon detectors and thermal detectors, such as thermocouples, thermistors and Golay detectors are the two usual types of IR detectors used in IR spectrometer. Thermal detectors measure the heating effects generated by IR radiation. For example, thermistor measures electrical resistance and thermocouple measures voltage at a junction of dissimilar metals. Furthermore, the thermal detectors exhibit a linear response even for a wide range of frequencies. However, they are not as sensitive and fast as photon detectors. The working of the photon detectors are based on interaction of IR radiation with a semiconductor material. As a result of the interaction, electrons from valence band excited to conduction band and hence, a small current or voltage is developed.

5.3.1 Fourier Transform Infrared (FTIR) Spectroscopy

Fourier transform spectrometers have superior speed and sensitivity as compared to dispersive instruments for most applications. In FTIR spectroscopy, all component frequencies are are viewed simultaneously. All molecules absorb IR light except monoatomic (e.g., He, Ne, Ar) and homopolar diatomic (e.g., H2, N2, O2) molecules. Molecules only absorb some IR radiations, whose frequencies affect the dipole moment of the molecule. The dipole moment of a molecule is due to the differences of charges in the electronic fields of its atoms. Molecules having only a dipole moment interact with infrared photons. The interaction can result the excitation of molecules to higher vibrational states. The electronic fields of atoms of homopolar diatomic molecules are equal and hence, they do not posses a dipole moment. No dipole moment associated with monoatomic molecules also because they have only one atom. Since monoatomic and homopolar diatomic molecules have no dipole moments, they do not absorb IR radiation.

As discussed above, each molecule only absorbs IR light of certain frequencies based on the characteristic of each molecule. Hence, the study of the absorption spectrum helps to identify the type of molecule (qualitative analysis) and the amount of molecule present in the sample (quantitative analysis).

The radiation source, interferometer and detector are the main components of an FTIR system. Figure 5.3 illustrates the working of a typical FTIR spectrometer. The IR radiation source of FTIR spectrometer contains a water cooling system to ensure better power and stability. An interferometer is used instead of monochromator, which divides incident beams and creates an optical path difference between the beams. The superposition of these beams produce interference pattern, which contain infrared spectral information. The interferometer used in FTIR spectrometer is the Michelson interferometer, which consists of a fixed mirror, a moving mirror and a beamsplitter. The two mirrors are mounted at right angles to each other. A semi-reflecting device is used as the beamsplitter. The beamsplitter splits the incident beam and transmits half of the beam to the fixed mirror and the rest is reflected to the moving mirror. The beams after reflection from the mirrors are superposed at the beamsplitter and an interference pattern is generated. The resulting beam is then passed through the sample, and finally collected by the detector.

Fig. 5.3: Schematic diagram of a typical FTIR Spectrometer

If the velocity of the moving mirror is constant, the intensity of beam collecting the detector changes sinusoidaly. The resulting interferogram output is shown in figure 5.3. The interferogram is a time domain spectrum that records the detector response changes versus time within the mirror scan. When the sample absorbs frequency of radiation, the amplitude of the sinusoidal wave will be reduced. Fourier transformation converts the time domain interferogram to frequency domain spectrum showing intensity versus frequency, the final IR spectrum. The FTIR spectroscopy has many advantages over dispersive method of infrared spectral analysis. This non destructive and fast technique gives precise measurement, which requires no external calibration.

5.4 Raman Spectroscopy

This technique is based on inelastic scattering of monochromatic light. In inelastic scattering, the frequency of light changes due to its interaction with a sample. Photons from a laser source are absorbed by the sample and re-emitted during de-excitation. The frequency of the emerging beam consists of both lower and higher frequencies in addition to the original frequency. This phenomenon is called the Raman Effect. The study of the change in frequencies gives information about vibrational, rotational and other low frequency transitions in molecules of solid, liquid or gaseous samples.

A typical Raman spectrometer consists of a source of light such as a laser, a double or triple monochromator and a signal processing unit composed of a detector, an amplifier and an output device. A sketch of a typical Raman spectrometer is shown in figure 5.4.

Fig. 5.4: Schematic diagram of a laser Raman system

A number of stages are involved in the acquisition of Raman spectrum. The sample on the sample chamber is illuminated with a laser light. A convex lens is used to focus light on the sample. Generally, liquid and solid samples are taken in a Pyrex capillary tube. The scattered light is collected and focused at the entrance slit of the monochromator using another lens. The width of the slit is adjusted for creating suitable spectral resolution. The light coming out through the exit slit of the monochromator is collected and focused on the surface of a detector. The detector converts this optical signal into an electrical signal and is further processed using detector electronics. Computer stores the output signal from the detector electronics for each predetermined frequency interval. A plot of the signal intensity versus wavenumber constitutes a Raman spectrum. The advances in solid state detectors replaced the conventional photomultiplier tube detectors with multichannel detectors (MCD). MCD based laser Raman spectrometers are more efficient and faster than the conventional Raman systems.

5.5 UV-VIS Spectroscopy

UV-Vis spectroscopic technique is useful to characterize the absorption, transmission, and reflectivity of materials. The principle of UV-Vis spectrometer is based on the fact that molecules of the sample absorb UV or visible light and results the excitation of outer electrons in the molecule. This absorption spectroscopy measures the absorption of light after it interacts with the sample. The measurements can be done either at a monochromatic wavelength or a wide spectral range. The wavelength of light absorbed by the sample is the characteristic of its chemical structure. Different spectral regions of the UV-Vis spectrum are absorbed by different types of molecules. As figure 5.1 shows, absorption of microwave radiation can result molecular rotational motion, and IR absorption results vibrational motion of molecules. But UV-Vis absorption can result transition of electrons to higher energy states. Each molecule undergoes electronic excitation following absorption of light. UV-visible light is adequate for molecules having conjugated electron systems (e.g., benzene absorbs light of wavelength 260 nm). The absorption spectrum shifts to the lower energy as the degree of conjugation increases (e.g., naphthalene absorbs light up to 300 nm and anthracene up to 400 nm). The study of the absorption spectra helps to identify atomic and molecular species, since they are characteristic of molecular structure.

The relation connecting the intensity of transmitted light through a solution of an absorbing material and the concentration of the material is given by Beer- Lambert law. According to the law:

- log (I/I0) = A = ¥¬ bc, (5.3)

where I0 and I are the intensity of incident and transmitted light respectively, A is the absorbance, ελ is the molar absorptivity (litre/mole/cm), b is the cell path length (cm) and c is the concentration of solution (moles/liter). The molar absorptivity represents the spectrum of the solution and is a function of wavelength. The value ελ is represented for a particular wavelength (e.g. ε532). Thus, the study of UV-Vis spectroscopy helps to measure the quantity chemical present in a solution.

UV-Vis Spectrophotometer consists of a UV light source, a monochromator and a detector. The monochromator works as a diffraction grating to dispense the beam of light into the various wavelengths. The detectors role is to record the intensity of the light which has been transmitted. Before the samples are run, a reference must be taken first to calibrates the spectra to screen out any spectral interference. In the case of liquid samples the solvent which has been used to dissolve the sample is used. However there is a criterion that the solvent should not absorb UV radiation in the same region as the sample being analysed. In the case of solid state UV-VIS spectroscopy the reference is normally KBr as it does not absorb radiation in the same region as most samples. This method is often used with samples that IR spectroscopy fails to identify.

5.6 Scanning Probe Microscopy (SPM)

SPM technique scans sample surfaces using a probe tip. The SPM has profound effects on many areas of science and technology. This technique is highly useful for the the investigation and manipulation of nanoscale materials. The probe of SPM is a narrow tip having a radius of curvature around 3-50 nm. The tip of the scanning probe is fixed on a flexible cantilever, which allows the probe tip to scan the surface profile. As the probe tip moves within the proximity of the sample surface, the forces of interaction between the surface and tip influence the movement of the cantilever.  The movements of the cantilever can be detected using suitable sensors. Various types of interactions can be studied with the help of SPM, depend on the mechanics of the probe. The SPM techniques can help to develop image clusters of individual atoms and molecules because they can operate even upto nanometers.

Based on interactions, SPMs are of various types such as, scanning tunneling microscopy (STM), atomic force microscopy (AFM) and near-field scanning optical microscopy (NSOM). STM measures electronic tunneling current, AFM measures interaction forces and NSOM measures local optical properties using near field effects (Fig.5.5). These characterization tools help to study the structural, mechanical, electrical and optical properties of materials in any environment.

Fig. 5.5: Schematic diagram of (a) a typical SPM and (b) the signals observed in STM, AFM and NSOM techniques.

5.6.1 Scanning Tunneling Microscopy (STM)

In 1981, Gerd K. Binnig and Heinrich Rohrer invented STM. They were awarded half of the 1986 Nobel Laureate in Physics for their work.  Three dimensional image of solid surface with atomic resolution was first obtained with STM. However, STMs can only be used to study electrically conducting samples. Mainly there are five scientific processes that the STM integrates to make atomic resolution. They are (i) quantum mechanical tunneling, (ii) controlled motion over small distances by piezoelectrics, (iii) negative feedback, (iv) vibration isolation and (v) electronic data collection. Each of these processes was known to the scientific community even before the invention of STM.

The electron tunneling principle was introduced by Giaever. Consider a system of two metals separated by an insulating thin film. When a suitable potential difference is applied across the metals, a current will start flowing because electrons can penetrate the potential barrier. It is possible to measure the tunneling current by spacing the two metals less than 10 nm. Vacuum tunneling together with lateral scanning was introduced by Binnig and his co-workers. The vacuum offers the ideal barrier for tunneling. The lateral scanning can help to image sample surfaces with wonderful resolution, even sufficient to image single atoms. Since the tunnelling current varies exponentially with the distance between the metal tip and the scanned surface, vertical resolution of the STM is very high. Usually, tunneling current reduces as the separation between the electrodes is increased. The lateral resolution depends on the sharpnes of the probe tips.

The tip of a typical STM is made from tungsten or Pt-Ir alloy that is attached to a piezo-drive. Three mutually perpendicular piezoelectric transducers: x- piezo, y- piezo and z- piezo are available with the piezo-drive, as shown in figure 5.6. When a p.d. is applied, piezoelectric transducer expands or contracts. The scanning waveform is formed between the x - y piezos and make the tip raster scan over the sample surface. The separation between the probe tip and the surface of the sample is kept very narrow (a fraction of nanometer) using the coarse positioner and the z- piezo. A suitable potential difference (0.1 V - 1 V) is applied across the tip and the sample. It can induce a tunneling current of about 0.1 nA to 1 nA. A negative feedback mechanism is employed to control the z- piezo, and to maintain a constant tunneling current. The voltage level on the z- piezo indicates the local height of the topography.

When the bias potential V between the tip and the sample is greater than zero (V > 0), electrons tunnel from the tip into the sample surface. Though, if V < 0, the electrons from the sample surface tunnel into the tip. By using a current amplifier, the tunneling current can be converted into a voltage. Compare this voltage with a reference value. The difference voltage is amplified to drive the z- piezo. The phase of the amplifier is chosen to provide a negative feedback. If tunneling current is higher than the reference value, the voltage applied to the z- piezo tends to withdraw the tip from the surface, and vice versa. Therefore an equilibrium z position is developed. As the tip scans over the xy plane, a two-dimensional array of equilibrium z positions will be obtained. This equilibrium position of the z-piezo represents a contour plot of the equal tunneling current. The contour plot can be displayed and stored in the computer.

Fig. 5.6: The schematic representation of a typical STM.

The topography of the surface is shown on a display monitor as shown in figure 5.7(a). In the figure, the bright and dark spots represent high z and low z values respectively. The scale bar represents the z values corresponding to the gray levels. Figure 5.7(b) shows contour plot along a given line, a quantitative representation of the topography. The unit for x and y is nanometer and that for z is picometer (10−12 m). The STM unit is made as rigid as possible to ensure vibration isolation, and thereby achieve atomic resolution.

Fig. 5.7: Model gray scale image and contour plot.

One disadvantage of the STM is that it requires conductive samples, so that at least some features of the specimen must show electrical conductivity to some extend. To circumvent this limitation, AFM was subsequently developed and it is used for studies of non-conducting samples.

5.6.2 Atomic Force Microscopy (AFM)

In 1986, Binning and Gerber discovered atomic force microscope to measure ultra small forces (< 1 μN), which present between the probe tip and the sample surface. This microscopic technique is an important tool for nanoscale imaging. It also helps to manipulate nanoscale materials. Besides these, AFM technique helps to study some physical and chemical properties of samples. One significant advantage is that AFM technique is suitable for studies of both electrically conducting and insulating samples.The resolution of the AFM is very high, of the order of fractions of a nanometer. The force of interaction between the tip and the sample surface depends on the separation between them and also nature of the sample surface.

Basic Principles

Fig. 5.8: Schematic sketch of a typical AFM.

A typical AFM consists of a cantilever with a probe tip, laser beam deflection system, a photodiode, PZT scanner, electronic feed back system and a detector (figure 5.8). The sharp tip attached to the end of the cantilever scans the surface of the specimen. The cantilever is made from Si or SiN. Its spring constant is about 0.01-100 N/m, and resonant frequency is about 5-300 kHz. When the tip of the probe moves near the surface of the specimen, the interaction force will cause a deflection of the cantilever. By Hooke's law, Force F = -kx, where k is the spring constant and x is the distance of the probe from the sample. Force distance curve is shown in figure 5.9.

Fig. 5.9: Force distance curve of an AFM.

The deflection of the cantilever is is measured using a laser beam deflection technique, as shown in figure 5.10. Beam of light from the laser source is reflected from the top surface of the cantilever. The reflected beam is made to incident on a split photodiode. Cantilever deflections are proportional to the difference signal VA−VB. It is possible to measure even sub-angstrom deflections. Hence, very small forces about tens of pico-Newtons can be measured. Beside this laser deflection method, methods like optical interferometry, capacitive sensing and piezoresistive cantilevers are used for the detection of cantilever deflection. However, these methods are not as sensitive as laser deflection method.

Fig. 5.10: Optical beam deflection system that detects cantilever motion in the AFM.

If the tip scans at a constant height, it collides with the sample surface and will be damaged. To overcome this difficulty, an electronic feedback system is used to adjust the tip-to-sample distance to maintain a steady force between them. The specimen to be scanned is placed on a piezoelectric tube. The tube moves the specimen along the z- axis for keeping a steady force. It also moves the sample along the x and y axes for scanning the surface of the specimen. Alternatively a tripod configuration of three piezo crystals can be used for scanning, as discussed in section 5.6.1. The advantage of this method is that it eliminates the distortion effects caused by a tube scanner. However in new systems, the probe tip is mounted on a vertical piezo scanner and the sample to be scanned is placed on a new piezo block. The resulting plot of the area s = f(x, y) represents the topography of the sample.

Imaging Modes

It is possible to operate AFM in different imaging modes, depending on the application or requirement. Static or contact mode and dynamic or non contact mode are the two basic modes of operation of AFM. In the first mode, the static tip deflection signal will be used as a feedback signal. Generally, a low stiffness cantilever is employed to enhance the deflection signal. But, within the proximity of the sample surface, attractive forces are strong as to cause the tip to collide the surface (figure 5.9). Inorder to maintain a steady deflection, the interaction force is kept steady during scanning.

In the non-contact mode, there is no direct contact between the tip of the cantilever and the sample surface. However, the cantilever oscillates at a frequency slightly greater than its resonance frequency. The amplitude of oscillation of the cantilever is typically a few nanometers. The characteristics of oscillation, such as amplitude, phase and resonance frequency are modified by tip-sample interaction forces. These modifications in oscillation with respect to the external reference oscillation give information about the characteristics of the specimen. Both the frequency modulation (FM) and amplitude modulation (AM) schemes are used in non-contact mode operation. In FM scheme, the frequency of oscillation will be modified and may provide information about tip-sample interactions. True atomic resolution will be obtained at ultra-high vacuum conditions. In AM, the amplitude or phase of oscillation will be modified and may provide necessary feedback signal for imaging. The modifications in the phase can help to explore various materials present on the sample surface. If the tip used in an AFM is sharper, the better the resolution. Carbon nanotubes are best suited for AFM tips because they are very strong and flexible, and also it may not be broken even though much force is exerted on it.

Besides imaging, force spectroscopy is another major application of AFM. Force spectroscopy helps to study and measure nanoscale contacts, atomic bonding, Casimir forces and Van der Waals forces.

(a) (b)

Fig. 5.11: AFM images of (a) gold nanoparticles and (b) sodium chloride

5.6.3 Near-Field Scanning Optical Microscopy (NSOM)

In 1920, Synge introduced near-field scanning optical microscopic technique with remarkable accuracy. This technique provides the highest lateral optical resolution. NSOM scans a small aperture (~100 nm) located very close to the specimen and detection of this light energy forms the image of the surface.  In the illumination mode of NSOM, a dielectric probe positioned at a distance d<<¬ from the surface illuminates the sample from above. Either the reflected or the transmitted beam of light is collected in the far field using detectors. A dielectric probe in the near field collects the transmitted light. In a typical NSOM, a tapered single-mode optical fibre tip scans over the sample surface using piezoelectric transducers. The tip is coated with a very narrow film of metals to reduce loss of light. This technique helps to obtain topographic and light intensity images simultaneously. NSOM optical fibre tip scans the surface of the sample, collects the total reflected intensity from each scan point and integrates it as one pixel to form the final image, as shown in figure 5.12. Figure 5.13 compares the resolution capabilities of scanning electron microscopy, conventional optical microscopy and NSOM.

Fig. 5.12: Schematic representation of NSOM.

Fig. 5.13: Resolution comparison of (a) SEM, (b) conventional optical microscopy,

(c) NSOM image and (d) NSOM image after Fourier deconvolution.

Besides optical fibre based NSOM, there is another type called apertureless NSOM. The apertureless NSOM is based on a sharp metallic tip that scans the sample. Technically it is possible to fabricate very sharp metallic tips (atomic size STM tips) as compared to optical fibre tips (~50 nm diameter). Thus, NSOM technique has the potential to achieve much better resolution.

5.7 Electron Microscopies

5.7.1 Transmission Electron Microscopy (TEM)

TEM is a high resolution microscopic technique capable of imaging fine details even up to a single column of atoms. Its resolution is much larger than that of the light microscopy. This high resolution of TEM is due to the small de Broglie wavelength of electrons.. TEMs are widely used in cancer research, virology, pollution, materials science, semiconductor nanoresearch and so on.

A typical TEM consists of a vacuum system, an electron gun which generates electron beam, a number of electromagnetic lenses and electrostatic plates. The electromagnetic lenses and electrostatic plates guide the electron beam as required. Imaging devices are subsequently used to create images which are formed from the interaction of the electrons transmitted through the ultra thin specimen. These images are magnified and focused on a fluorescent screen or a charge coupled device (CCD) camera.


In 1923, de Broglie introduced the concept of matter waves, which states that all particles have an associated wavelength linked to their momentum. The de Broglie wave length ¬ = [h/mv], (5.4)

where m is the relativistic mass, v the relativistic velocity and h the Plank's constant. This equation is called de Broglie wave equation. Hans Bush (1927) found that it is possible to focus an electron beam using a magnetic coil as a glass lens focus light. In 1931, Ernst Ruska and Max Knoll recorded the first TEM image. Reinhold Rudenberg of Siemens Company took the patent of an electrostatic lens electron microscope during the same year.

The wavelength of electrons (¬e) can be measured by equating the de Broglie wave equation to the kinetic energy of an electron. A relativistic correction is applied because the velocity of electron approaches the speed of light c.

The wavelength, ¬e » [h/-{2m0E (1+E/2m0c2)}]. (5.5)

The emitted electrons from the gun are accelerated by an electric potential of about 100-1000 kV and their velocity approaches to the speed of light (0.6-0.9c). These high speed electrons are focused on the sample surface using electrostatic and electromagnetic lenses. The transmitted electron beam is used to develop images. The magnetic lens aberrations limit the TEM resolution to the Å order. This resolution of the TEM is suitable for material imaging and structure determination at the atomic level.

Source Formation

The electron gun in a TEM generates electrons by thermionic or field electron emission. Schematic layout of optical components in a typical TEM is shown in figure 5.14. The manipulation and focussing of the electron beam is made using electromagnetic and electrostatic effects. When electron enters the magnetic field of electromagnets, they start moving according to right hand rule. It is possible to form electromagnetic lens of variable focusing power. The electrostatic fields can deflect the electrons through desired angle. These two effects together with an electron imaging system can help to control the electron beam path. Furthermore, it is possible to modify the optical configuration of a TEM.

Fig. 5.14: Layout of optical components in a basic TEM


A typical TEM mainly consists of condensor lenses, objective lenses and projector lenses. The condensor lenses form primary beam, and the beam is focused into the sample surface using objective lenses. While, projector lenses expand the beam onto the fluorescent screen.

Fig. 5.15: Transmission electron micrographs of CdSe quantum dots.


The display unit consists of a phosphor screen and an image recording system. The phosphor screen is made of fine zinc sulphide particles (10-100 nm) for direct observation. The image recording system is a film or doped YAG screen coupled CCDs. It is possible to rotate sample by a desired angle, and thus to take different images of a sample at different angles (in 10 increments). These images can be used to construct a 3D image of the sample. TEM images of CdSe quantum dots are shown in figure 5.15.

5.7.2 High-Resolution Transmission Electron Microscopy (HRTEM)

HETEM is a powerful tool for studying the structure of particles, interfaces and crystal defects. This microscopic technique images crystallographic structure of materials with atomic scale precision. HRTEM provides highest resolution about 0.08 nm with microscopes. Because of this high resolution, this unique tool help to study the properties of nanocrystalline materials like semiconductors and metals. The phase-contrast imaging is the basis of image formation in HRTEM. In this type of imaging, the contrast may not be interpretable spontaneously because the image is affected by aberrations of imaging lenses.

Basic Principle

A thin sample of crystal is mounted such that a low index direction is at right angles to the incident electron beam direction. The primary beam of electrons will be diffracted by all lattice planes parallel to the electron beam. The diffracted and the primary beam are made incident on the objective lens. Their interference result a back transformation and produces an enlarged image of the periodic potential. This image can be further magnified by the electron optical system and will be formed on the screen. This imaging process is called phase-contrast imaging and the image formed is an indirect depiction of the crystallographic structure of the specimen. The phase of the electron wave carries information about the structure of samples and generates contrast in the image. Hence, the name phase-contrast imaging. The principle of working is schematically illustrated in figure 5.16.

Fig. 5. 16: Schematic diagram of the electron beam column in HRTEM.

The interaction of the electron wave with the crystallographic structure of the sample gives a qualitative information about the structure of the specimen.. Each imaging electron interacts independently with the sample. The electron wave can be considered as a plane wave and is made incident on the sample surface. When it enters the specimen, the electron beam is attracted by the positive potentials of the atomic cores, and directs it along the atomic columns of the crystallographic lattice. The interaction between the electron waves in different atomic columns results Bragg diffraction. HRTEM image of a CdSe nanocrystal is shown in figure 5.17.

Fig. 5.17: HRTEM image of a CdSe nanocrystal.

5.7.3 Scanning Electron Microscopy (SEM)

SEM is widely used for the characterization of nanomaterials and nanostructures. Its resolution is nearly a few nanometers. The magnification of SEM can be adjusted from 10 to around 300,000. It is interesting that SEM provides the chemical composition information near the surface of the sample in addition to topographical information. High energy electron beam from an electron gun is focused onto the surface of the specimen to produce a number of signals, as shown in figure 5.18. The signals that are derived from the interactions between electron and sample could provide information about the sample, such as chemical composition, crystalline structure and external morphology (texture).

Schematic diagram of a scanning electron microscope is shown in figure 5.18. A SEM composed of a source of electron, electromagnetic lenses, vacuum system, detectors and a display device. In a SEM, a beam of electrons having energy ranging from a few hundred eV to 50 KeV is made incident on the sample surface. A verity of interactions occurs that result in the emission of number of electrons and photons from the sample. The emitted electrons consist of secondary electrons, backscattered electrons and diffracted backscattered electrons. The SEM employs secondary electrons and backscattered electrons for imaging samples. Surface morphology and topography of the sample can be obtained from the secondary electrons. Crystal structures and orientation of materials will be derived from the diffracted backscattered electrons. The photons emitted during the interaction may consist of characteristic X-rays, continuum X-rays and visible light. When a beam of electrons is incident on the specimen, electrons in discrete orbitals will be excited. During the process of deexcitation, the excited electrons are return to lower energy states and as result, characteristic X-rays of fixed wave length will be produced. These X-rays are suitable for elemental analysis. The scanning electron microscopic technique is a non-destructive method because the X-rays produced during the interaction do not result any volume loss of the specimen. Hence, the same materials can be analysed repeatedly.

The resolving power, R, of an instrument is defined as:

R =¬/(2NA), (5.6 )

where ¬ is the wavelength of electrons used and NA is the numerical aperture, which is engraved on each objective and condenser lens system, and a measure of the electron gathering ability of the objective, or the electron providing ability of the condenser.

Fig. 5.18: Schematic diagram of a scanning electron microscope.

The electron gun emits a narrow beam of electrons and these electrons are focused using magnetic lenses. A coil of copper wire produces a magnetic field that is shaped into a suitable geometry to create the electromagnetic lensing action, similar to that of an optical lens. If an electron of charge q moves with a velocity v through a magnetic field B, it experiences a radial force F inward, called magnetic Lorenz force. Magnetic Lorenz force F = q(v x B). The focal length of the electromagnetic lens depends on the gun voltage and the amount of current through the coil. The velocity of the electron beam depends on gun voltage and the flux density depends on the amount of current through the coil. Therefore, it is possible to control the focal length of the electromagnetic lens by controlling the current through the coil. As current increases, the radial force experienced by the beam increses and hence, the focal length of the electromagnetic lens will be reduced.


Fig. 5.19: SEM images of nanorod arrays

In SEM, electron beam scan the surface of sample. There are two coils for scanning, one for for raster and the other for deflection. These coils are positioned in the bore of the objective lens cage (Fig. 5.18) and help the electron beam to scan over a square area on the sample surface. SEM images of nanorod arrays are shown in figure 5.19.