Lecture 12 - Nanotechnology PDF

🔬 Lecture 12 - Nanotechnology Introduction Nanometers : 10^(-9) meter Nanotechnology involves individual manipulation of single molecules or even atoms. Building components atom by atom or molecule by molecule to create materials...

🔬 Lecture 12 - Nanotechnology Introduction Nanometers : 10^(-9) meter Nanotechnology involves individual manipulation of single molecules or even atoms. Building components atom by atom or molecule by molecule to create materials with novel and vastly imroved properties = the original goal of nanotechnology. Now it also expanded to include any structure so tiny that their study and maniuplation was impossible until recently. Quantum effects at nanoscale are covered. → Internal biological components of cells are on the same scale as those studied in nanotech. Most nanotechnology is in fact molecular biology viewed from the perspective of material science. ⇒ The main practical objectives of nanobiotechnology are using biological components to achieve nanoscale tasks. Some of these tasks are nonbiological and have applications in such areas as electronics and computing, whereas others are applicable to biology or medicine. Visualization at the nano-scale Scanning probe microscope : opened up the field of nanotechnology by allowing visualization of individual atoms and molecules. Concept : measuring some property (electrical resistance, magnetism, temperature, light absorption) with a tip positioned extremely close to the sample. 1. The microscope raster-scans the probe over the sample while measuring the property of interest. 2. The data are displayed as raster image (similar to television screen). Unlike traditional miscroscopes : scanned probe systems do not rely on lenses : resolution is not limited by diffraction anymore, but by the size of the probe. Some scanning probe instruments proceed simultaneously : visualization + alteration of the sample. The first scanning probe instruments : 1. scanning tunneling microscope (STM) developed at IBM - measure electrical resistance 2. Atomic force microscope (AFM) : measure the force between probe tip and the sample. Scanning tunneling microscopy Metal tip comes close to a conducting surface : electron can tunnel in either direction. The probability of tunneling is directly related to the distance between the tip an the sample. By keeping the current constant and measuring the height of the tip accordingly, we can map the surface contours of the Lecture 12 - Nanotechnology 1  sample. The tip can detect each individual atom on the surface being studied and renders therefore high resolution. We can also remove and rearrange atoms by using STM. Weakness of STM in biology : requires a conducting surface in contrast to ATM. Atomic force microscopy Concept : a sharp probe moves over the surface of the sample and bends in response to the force between the tip and the sample. The movement of probe tip performs a raster scan. The resulting topographical image is displayed on-screen;. The positioning and movement of the tip (or sample), during scanning, is done by a precise positioning device made from piezoelectric ceramics (change shape in response to an applied voltage). The AFM probe is a tip at the end of a cantilever. As the cantilever bends because of the force (repulsive molecular interaction between the tip and the sample) , a laser monitors its displacement by emitting a beam of light that is collected on a position-sensitive detector, after being reflected on a photodiode on top of the cantilever. The difference in two consecutive signals gives information about the the bending and thus the displacement of the tip. → It can also be used to move a single atoms (like STM) but only since 2003. → It helps visualize polymeric biological molecule such as DNA or cellulose, see individual monomers and at high resolution event the atom composition. Weighing single bacteria and virus particles In addition to visualize microorganism, we can weight them. Lecture 12 - Nanotechnology 2  With a laser we can measure the oscillation frequency of a micrometer long cantilever (scaled down). Adding a bacteria, virus, etc can modify this oscillation frequency : the mass of the organism can be deduced. An antibody can be used to immobilize the bacteria or virus on the cantilever. → An array of cantilevers (rather than a single cantilever) would be used when counting particles for quantification. Nanoparticles and their uses Nanoparticles : 1. submicron scale : 5 to 100 nm in size 2. spherical usually, but also rod, plates or other shapes are used 3. solid or hollow 4. composed of variety of materials distributed in several discrete layers with separate functions : a. The central functional layer : for useful optical and magnetic behavior - most popular is fluorescence b. The protective layer : shields the functional layer from chemical damage by air, water or cell component + conversely protect the cell from the chemical components of the functional layer. c. The outer layer/layers : allow the nanoparticles to be “biocompatible” ⇒ water solubility (hydrophobic layer) and specific recognition (chemical binding molecules attached). Lecture 12 - Nanotechnology 3  Uses of nanoparticles in biological arena (analytical and clinical) : (a) Fluorescent labeling and optical coding (b) Detection of pathogenic microorganisms and/or specific proteins (c) Purification and manipulation of biological components (d) Delivery of pharmaceuticals and/or genes (e) Tumor destruction by chemical or thermal means (f) Contrast enhancement in magnetic resonance imaging (MRI) Nanoparticles for labeling Luminescent CdSe nanorods = used for fluorescent labeling. They absorb at 550 nm and emit strongly at 590 nm. 1. Core of luminous cadmium selenide (CdSe) 2. Outer shell of zinc sulfide, wurtzite (ZnS) to protect the core against oxidation. 3. Outside layer = silica layer allow the coupling of hydrophilic groups (amines, etc) to make the nanorods water soluble. These outer chemical groups can also allow the attachement of nanorods to proteins. Used to mimic native tubilin found in the cell. Tubilin monomers form a cylindrical protein structure called microtubules. Nanorods can insert them in the assembly process = synthetic fluorescent microtubules. Why use complex multilayered nanostructure instead of simple fluorescent dye ? Lecture 12 - Nanotechnology 4  In contrast to typical dye, the absorption peaks are broader and the emission peak are narrower : it reduces the photobleaching of nanocrystals. Long-term irridation and monitoring is possible. Nanocrystals have high brightness (high molar absorptivity x quantum yield). The emission maximum is related to the size of the nanocrystals - thus we can tune the emitted wavelength by changing the size. Nanoparticles can target specific tissues by adding appropriate antibodies or receptor proteins to the nanoparticle surface. Fluorescent nanoparticles are known as quantum dots - they are commercially available and more versatile than fluorescent dye that attaches to molecule. We can label PCR primers with quantum dots to fluorescently label PCR product = quantum dot PCR Quantum size effect and nanocrystal colors The quantum dots are in fact semiconductors (SMC) that are small enough to show quantum effect. Semiconductors are substances that conduct electricity only under some conditions but not others. N-type semiconductors (as electric wires) : current consists of negatively charged electrons | P-type semiconductors : current consists of holes (absence of an electron from an atom). A hole can “move” from one atom to another. Electrons and holes may combine and cancel out - this produces energy. Conversely, energy absorbed by certain SMC can generate an electron-hole pair whose element can move in diff. directions. Nano particle labels can be made in diff. emission wavelength : in UV, visible spectrum, near infrared. depending on the semiconductor material depending on the quantum size effect : the smaller the nanoparticles, the more energy is needed (to produce the electron-hole pair and confine it - quantum confinement) and the shorter the wavelength (the higher energy) it emits. Light is emitted when the hole and an electron recombine. Nanoparticles for delivery of drugs, DNA, or RNA Lecture 12 - Nanotechnology 5  Nanoparticles can target specific tissues and deliver a variety of biologically active molecules such as pharmaceuticals (drugs, etc) and genetic engineering constructs (DNA, RNA) : Biomolecule itseft - DNA can form nanoparticles (we add positively charged molecules to neutralize it and can add also additional molecules to promote the selectivity of cells or tissues). Alternatively = hollow biocompatible nanoparticle (nanoshells) can carry other smaller molecules. Ex. Chitosan nanocarries (matrix system or core-shell system) which interacts strongly with mucosal surfaces. Ex. combination of nanoshells to carry interfering RNA Nanoparticles in cancer therapy Nanoparticles may be used to kill cancer cells by : 1. localized heating (using magnetic core under magnetic field or metal nanoshell tuned to absorb radiant near infrared and thus heat surrounding tissue) or 2. by local generation of a toxic product such as singlet oxygen. The near- infrared laser excites the dyes attached to the nanoparticle. Energy transfer to photosensitizers by fluorescent resonance energy transfer (FRET) results in conversion of normal (triplet) oxygen to singlet oxygen 3. Another approach to cancer therapy is using nanoparticles to regulate blood vessel formation (gold nanoparticles with short surface peptides that bind to receptor of specific cells taking part in angiogenesis to prevent blood capillary formation). Assembly of nanocrystals by microorganisms Bacteria can accumulate a variety of metallic elements and modify them chemically : oxidation and reduction. Ex: E.coli when exposed to cadmium chloride (CdCl2) and sodium sulfide (Na2S03), it precipitates cadmium sulfide (CdS) as particle in 2-5 nm size. Hence, bacteria can “biosynthesize” semiconductor nanocrystals. Phage display can be used to select peptides capable of binding to nanocrystals such as ZnS or CdS and thus creating semiconductor nanowire : because the bacteriophage M13 mostly used is filamentous, the binding of the nano-crystals on its hybrid surface proteins forms a “semiconductor nanowire”. Lecture 12 - Nanotechnology 6  Nanotubes Carbon nanotubes : cylinders made of pure carbon with diameter 1-50 nm and up to 10 mm long. Other pure elemental carbon is found either as diamond (each carbon is covalently linked to four others - forming a strong lattice) or graphite (flat sheets of carbon that form hexagonal pattern - each carbon has 3 covalent bonds - the sheets can slide sideways over each other because there is no covalent linkages between atoms in different sheets). To form nanotube : single sheet of graphite is rolled into a cylinder. They can act as metallic conductor or semiconductor. Diameter and torsion can be tuned. Single-walled carbon nanotubes are especially used in biotechnology because it enters very readily. Attaching useful molecules to them (enzyme, antibody, etc) is the critical issue : the surface of carbon nanotube is hydrophobic ! Method 1 : adding nonionic detergent with a hydrophobic portion binding to the hydrophobic surface and hydrophilic region bidning to protein. Method 2 : chemical reagents reacting with the carbon on the surface, generating side chains carrying reactive functional groups. Proteins can then be linked covalently by reaction with these. Possible application of carbon nanotube in biotechnology and medicine : 1. Imaging : carbon nanotube show intrinsic luminescence in the near infrared - NIRM (near infra-red microscopy) can detect it. 2. Electrochemical sensors : when the surface-binding chemical groups of nanotubes interact with other molecules, it can change its electrical properties = electrical signal can be generated upon detection of a Lecture 12 - Nanotechnology 7  target molecule. 3. Photothermal killing of cancer cells : it absorbs near infrared radiation and generate local heating causing cell death. Nanotubes needs to carry molecules specifically targeting cancer cells. 4. Drug delivery : linker molecule between the surface and the drug - the drug is attached and not encapsulated ! Ex. Paclitaxel (PTX) delivery. 5. Tissue regeneration : nanotubes with appropriate chemical groups can act as scaffold for regenerating tissues. Antibacterial nanocarpets Nanotubes may be assembled to create surfaces (nanocarpets) that are antibacterial or act as biosensors. Detection of viruses by nanowires Nanowire sensors are capable of detecting specific individual viruses. Binding of a virus particle changes the conductance of the nanowire. Ion channel nanosensors Somewhat more complex than nanotubes and nanowires are nanoscale ion channels that are assembled into membranes. These channels are designed so that they can be controlled to permit the movement of ions under only certain conditions. The ion flow generates an electrical current that is detected, amplified, and displayed by appropriate electronic apparatus. → used to detect variety of target molecules. Nanoengineering of DNA The goal : using DNA merely as a structural material rather than manipulating genetic information. double helix is a convenient structural module natural baise-pairing allows the linkage of separate DNA molecules together the requirement to create 3D structure is branched DNA : mixing up four carefully designed single strands with different sequences can generate cross-shaped DNA. Each strand base-pairs with two other strands over half its length. If sticky ends are included in the initial strands : we can link the crosses together in 2D matrix. Nicks can be sealed with DNA ligase. Lecture 12 - Nanotechnology 8  Principles of branching can be extended to 3D : we can build cubical DNA lattices. Use : To build nanoscale framework - which can be used for the assembly of other components (metallic nanowires, nanocricuit, etc) Drawback : junctions are flexible (not a 90°) in cross-shaped DNA molecules and theri 3D counterparts. A rigid DNA component has been made by using double-crossover (DX) DNA molecules : DNA origami Building nanostructures by assembling multiple different DNA molecules becomes difficult beyond a certain level of complexity. Therefore DNA origami approach simplifies building DNA nanostructures by using one very long DNA strand and folding it up to form a scaffold. Staple strands are added in excess to help folding. They bind to specific sites along the longer scaffold strand to drive folding. In this approach, there is no need to strictly control the ratio of different DNA strands as for traditional DNA folding (cross-shaped DNA, etc). Assembly is much faster and yields are higher in this approach. Can rely on computer-aided design : to specify the required DNA sequence for a specific desired form. Lecture 12 - Nanotechnology 9 Lecture 12 - Nanotechnology 10