APPLICATION OF SCANNING ELECTRON MICROSCOPY AND TRANSMISSION ELECTRON MICROSCOPY IN NANOTECHNOLOGY

GUIDED BY: Dr.SAMEER PATEL PRESENTED BY : NEHAL LAKUM ID-10MPHPT006 PCETICAL TECH.

Points to be covered

Introduction Instrumentation of SEM Diff between SEM & TEM Application of SEM & TEM in Nanotechnology

TYPES OF ELECTRONE IN ELECTRONIC IMAGING:

1. Secondary electrons (SEs):A collision of electrons from primary electron beam with the surface of the specimen results in a detachment of it so called SEs. 2. Backscattered electrons (BSEs):these are primary electrons that have been reflected from the specimen 4. Transmitted electrons: that are transmitted from specimen 5. Absorbed electrons: that are absorbed by specimen.

Signals arising from the specimens surface on the primary electronic beam

Principle:

The basic principle of an electron microscope is based on the light microscope except that electrons are used instead of light.

Instrumentation

The electrons are emitted by the cathode from a filament, normally a heated tungsten wire. The emitted electron current is controlled by a Wehnelt cylinder having a negative polarity against the cathode. The electrons are picked up by the anode after being accelerated by the accelerating voltage of approximately 125 kV. The cathode assembly and the anode are arranged so as to produce a crossover of the electron beam between the components. Through the bore hole of the anode, the electron beam enters two electromagnetic condenser lenses which reduce the crossover. After passing the aperture the electron beam is focused by the objective lens so that the focal spot is imaged on the specimen surface.
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The specimen is scanned point by point by the electron beam as shown in Fig. SEs that are emitted from the specimens surface are collected by the detector and undergo a video processing leading to the image formed on the video screen.

The electron beam is moved on the surface of the specimen by an electromagnetic deflection system that is integrated in the objective lens moving the beam in a raster over the specimen as mentioned above. The deflection system consists of two sets of crossed saddle coils for deflection in X and Y direction. The saddle coils produce distortion-free images at lowest magnifications and permit large deflection angles.
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The SE detector is mounted to the side of the microscope chamber. PEs liberate secondary ones (SEs) from the specimens surface, which are caught by the collector having a positive potential for the detection of SE. The electrons pass through the grid and move toward the scintillator. These electrons strike the scintillator, where they generate photons, which are guided out of the detector chamber through a light guide to the photomultiplier The photomultiplier converts the light current by amplification again into an electron current that presents the video signal at the output of the subsequent preamplifier. If the collector is at a negative potential, only BSEs can strike the scintillator and a BSE image is produced in this mode. 9

Scanig electron micropscope (SEM)

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Indusrial view of TEM

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Difference between SEM&TEM

The TEM differs from the SEM, as its electron beam hits the sample, the electron beam is transmitted or passes through the sample and several more lenses to a phosphorus screen.The image is generated. Darker areas represent areas of the sample where fewer electrons where transmitted through and lighter areas represent where more electrons where transmitted through It. The increased resolution results from the shorter wavelength of the electron beam, increasing resolution in the to a theoretical limit of 0.2 nm. The magnifications reached by TEMs are commonly over 100,000X.

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SEM

TEM

Electrongun,2lenses,1 aperture, sample and movable stage Less resolution then tem
Sample Almost any clean form: solid.Big, thick samples Most useful for: Fracture, wear or corrosion surfaces, powder polished & etched microstructures, IC chips, chemical segregation.

Electron gun 4lenses, 2apertures,sample movablestage. More resolution then sem

Foil or powders < 1000 Ang. thick. or surface replicas Selected area diffraction, imaging of dislocations, tiny precipitates, grain Boundaries and other defect structures in solids.
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Application of SEM AND TEM in Nanotechnology

In nanocapsules Identification of nanoporous membrane In nanoparticales & nanocrystales In nanosphere In metal complexes In cationic cotton fiber IN liposomes & Niosomes In nanosuspension In nanoemulsion Nanopellets Nanocreams In solid dispersion & inclusion complex.

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1)

IN NANOCAPSULES:

Nanocapsules have been made for many years following the example of nature, using molecules called phospholipids, which are hydrophobic (water-repellant) on one end and hydrophilic (water-loving) on the other. When these molecules are placed in an aqueous environment, they can spontaneously form capsules in which the hydrophobic portions are inside,protecting them from contact with paper.Nanocapsules are between 130 to 600 nm in diameter The capsules act like sponges, soaking up and holding the cream inside until the outer shell dissolves under your skin, rather than sitting on the top layer. Therefore, the nanotechnology enhanced creams are affecting the lower skin layers.
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nanocapsule

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Labs at the French National Center for Scientific Research and LOreal develope nanocapsules with biodegradable polymeric shells small enough to pass through the first layers of the skin. The skins natural enzymes dissolve the nanocapsule surface and release the active ingredients to lower layers of the skin. To gain the most benefits, active ingredients (vitamin A, retinol and beta-carotene) should get to the deeper layers of the skin.

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2)

IDENTIFY NANOPOROUS BIOLOGICAL MATERIAL

Nanoporous are materials all about holes that are less than 100 nm. They abound in the natural world, both in biological systems and in natural minerals. For example, the walls of our cells are nanoporous membranes and thepetroleum industry has been using naturally nanoporous materials called zeloites as catalysts for decades, though the majority used now a synthetic. Some of the most interesting applications for nanoporous membranes come from the ability of A nanopores of certain sizes to let some substances pass and others not, or to force molecules like DNA to pass through one at a time.Controlling the size of these pores accurately is one of the technological challenges faced in making these materials,Nanoporous membrane

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Nanoporous membrane

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3) IN NANOPARTICALS & NANOCRYSTALS

Nanoparticles and nanocrystalline materials are already commercialized as antimicrobial and antifugal agents. The health care industry needs for improved against bacteria in the face of growing antibiotic resistance. Silver has antibiotic properties and is being used to made into crystalline nanoparticles, which increase solubility and potency.There are a development which consists of nanodroplets of oil that can destroy bacterial spores, viruses and funguses. Another potential weapon against antibiotic-resistant bacteria lies in self-assembling nanotubes made of peptides.
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Nanoparticales of 1) silver

2) gold
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Crystal of pb

This image of a lattice crystal was captured by Cornell researchers using a scanning transmission electron microscope (STEM) at IBM. The yellow circles in the center of each pear-shaped molecule represent the stronger signal produced by a large atom; the red portions that make up the top of each pear shape show the weaker signal of the smaller atoms. The image allows researchers to see the orientation of the individual atoms within a crystal for the first time, thus giving researchers a vital tool for predicting the crystal's properties. A model of the molecular structure is superimposed

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A reconstructed image of a 9 nm diameter CdS quantum dot along the crystal's hexagonal axis

Cds crystal

diamond crystal

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4) IN NANOSPHERE IDENTIFICATION

nanosphere

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5) IN METAL COMPLEXES
TEM is very useful in identifying various complexes of metal.

TEM images of 20 wt% platinum-xGnP at (a) low magnification, (b) high magnification

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(TEM) image of a calcium silicate hydrate

polymer nanosize

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6) IDENTIFY THE MORPHOLOGY

Scanning electron microscope (SEM) images of particles of various sizes, shapes, and compositions prepared (a) hydrogel rods containing antisense oligonucleotide; (b) crosslinked degradable matrix cubes containing doxorubicin HCl; (c) abraxane harvested onto medical adhesive; (d) insulin particles harvested onto a medical adhesive; (e) hydrogel boomerangs containing 15 wt% iron oxide; (f) hydrogel cylinders containing 10 wt% Omniscan Transmission electron microscopy (TEM) image showing cell internalization of 150 450 nm (top) or 200 200 nm (bottom) cylindrical particles
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7) Identify cationic cotton fiber

Transmission Electron Microscopy Image of a cationic cotton fiber conformally coated with 20 sequential nanolayers of polystyrene sulfonate and poly(Allyl amine Hydrochloride). The thickness and composition of each layer controls the transport of chemicals through the modified textile without affecting its comfort or mechanical properties.

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Transmission Electron Microscopy Image of a cotton fiber coated with gold nanoparticles The nanoparticles are assembled on the surface of the fiber by controlling the electrostatic interactions between the cationic cotton and the anionic nanoparticles.

Potential applications include catalytic mantles as well as platform for biological sensors. Due to the high surface coverage on the surface of the fiber plasmonic effects are observed hence creating coloration without the use of dyes or pigments.

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TEM image of coated gold fiber

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8) LIPOSOMES BY Transmission electron microscopy

studies also indicated that the particle size, vesicle shape and lamellarity of liposomes may be different due to the process of preparation. Very often the preparations are metastable. That means the state of free enthalpy is not in an equilibrium with the environment. As a result the vesicles change their lamellarity, size, size distribution, and shape with time. For example, small vesicles tend to form larger ones and large vesicles smaller ones. This can be identified by TEM.
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The transmission electron micrographs in different fields (a-d) of the liposomes prepared by thin film method

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9) In nanosuspension

Nanosuspensions as aqueous formulations need to be preserved.This can be evaluated by SEM&TEM. Different size of particles can be determined.

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nanosuspension

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10) nanoemulsion

NB-002, a Novel Nanoemulsion with Broad Antifungal Activity against Dermatophytes, Other Filamentous Fungi, and Candida albicans For scanning electron microscopy, samples (450 l) obtained at different time points during the time-kill study with NB-002 were mixed with 113 l of fixative (10% aqueous solution of glutaraldehyde). The mixtures were vortexed and placed at 4C for at least 18 h. And mounted on scanning electron microscopy stubs by using a mixture of colloidal graphite and Duco cement. Samples for electron microscopy were sputter coated with gold by using a Polaron sputter coater (Quorum Technologies, United Kingdom), examined on an Amray 38 1910 FE scanning Electron microscopy

SEM of NBD030 after treatment with NB-002 for 1 h at room temperature. (A) no treatment or treatment with vehicle; (B) treatment with 100 g NB-002; inset, higher magnification to show the differential size of the nanoemulsion droplets (white arrow, NB-002) compared to the various sizes of the extrusions (blebs); (C) control microconidia (no treatment or treatment with vehicle; white arrow, microconidial spore); (D) treatment of microconidial spores with 12.5 g 39 NB-002, with white arrows indicating broken spores.

12) In nanopallets

Graphene Platelet Nanopowder Appearance: Black powder Thickness: 6-8 nm Average Particle Diameter: 15 micron Surface Area: 120-150 m2/g Carbon: 99.5+% Electrical Conductivy. identified by tem images

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nanopellets
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13) In nanocreams:

PhytoCellTec stem cell cream is formulated with nanotechnology that allows a higher encapsulation of the PhytoCellTec stem cell cream active ingredients in a smaller molecule. This minimizes trans epidermal water loss, enhances new stem cell production and allows better penetration of our PhytoCellTec stem cell cream. StemCellin's remarkable stem cell cream is delivered deep into your skin by the ultrapenetrating nano-emulsion that is released .
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Marketed nanocreames

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14) in inclusion complex:

Tamoxifen-2-hydroxylpropyl--cyclodextrin-aggregated Nanoassembly for Nonbreast Estrogen-receptor-positive Cancer Therapy

Tamoxifen (Tam) is used for the treatment and prevention of estrogen-receptor-positive human breast and other cancers.

Cyclodextrins are used to form inclusion complexes with a variety of drug molecules and, therefore, primarily resulting in improvements of dissolution and bioavailability. This, in turn, enhances solubility and improves chemical and physical stability.
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The transmission electron microscopy of TamHPCD indicated the transition of Tam from crystalline to amorphous form on addition of HPCD. Transmission-electron microscopy studies of HPCD and the Tam-HPCD complex revealed the formation of aggregated nanoassembly at 60-180 nm The Tam-HPCD nanoassembly entered the cell owing to enhanced permeability
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TEM image of cyclodexrin inclusion complex

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15) In solid dispersion

Therapeutic potential of curcumin could often be limited by its poor solubility, bioavailability, and photostability. To overcome these drawbacks, efficacious formulations of curcumin, including nanocrystal solid dispersion (CSD-Cur), amorphous solid dispersion (ASD-Cur), and nanoemulsion (NE-Cur), were designed with the aim of improving physicochemical and pharmacokinetic properties. Physicochemical properties of the prepared formulations were characterized by scanning/transmission electron microscope for morphological analysis. Significant improvement in pharmacokinetic behavior was observed in the newly developed formulations.
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TEM images of a) amorphous curcumin b) crystalline curcumine

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Conclusion

SEM & TEM are very useful in identification of nano partical size,shape & structure. It is a useful tool to characterize raw materials. Intermediateslike powder mixtures, granules, and other bulk materials could be visualized by SEM & TEM. Thus it is very useful in nanotechnology

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REFERENCEs

1) Principles of instrumental analysis by skoog and Holler,5th edition, pg no:539 2). Paulus Seitavuopio, Jyrki Heinmki, Jukka Rantanen, Jouko Yliruusi. Journal of American Association of Pharmaceutical Scientists .Volume 07, Issue 02 2000 3). R. Ficarra P. Ficarra, M. R. Di Bella, D. Raneri S. Tommasini M. L. Calabr M. C. Gamberini and C. Rustichelli, Journal of Pharmaceutical and Biomedical AnalysisVolume 23, Issue 1, August 2000, Pages 33-40. 4). Almira I. BlazekWelsh and David G. Rhodes, Journal of pharmaceutical research , Volume 18, Number 5 / May, 2001, pageno:652
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5). Jingyue Liu Journal of Electron Microscopy August 25, 2005

pages 251-278;

6). Guo Y, Gonzalez S, Guadalupe AR. Journal of electron microscopy. 1999 7). Encyclopedia of pharmaceutical technology , 3rd Edition, Informa healthcare, Vol. 4. 2007 ,Pages 3226- 3254 8). Watt, I.M. Microscopy with light and electrons. In The Principle and Practice of Electron Microscopy, 2nd Ed.; Cambridge University Press: New York, 1997; 810.

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websites

1 Jmicro oxfordjournal.org 2. www.nanotechnology.org 3. http://www.photometrics.net/sem.html 4. http://www.purdue.edu/REM/rs/graphics/sem2.gif 5. http://www.unl.edu/CMRAcfem/semoptic.htm 7. www.pharmainfo.net/coating-articles 8. www.pharmainfo.net/reviews/nanoparticles

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THANK YOU

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