Nanoparticles Flashcards
(16 cards)
Characterising nanoscale systems - three main categories
- Spectroscopic methods
- Imaging/microscopy methods
- Mass spectrometry methods
Characterising nanoscale systems
- Spectroscopic methods
- UV-Vis
- Dynamic Light Scattering (DLS)
Characterising nanoscale systems
- Spectroscopic methods
- UV/Vis
UV-Vis: Certain metallic/semi-conducting nanoparticles called quantum dots are coloured (absorb in the visible) due to surface plasom resonance that in turn depends on size or diameter of the particle. → Possible to use UV-Vis to estimate the size of these particles
Characterising nanoscale systems
- Spectroscopic methods
- DLS
Dynamic light scattering (DLS): The intensity and time-dependency of the Rayleigh/Mie scattering of light from particles in the range of a few nm to microns (≈ 1-5000 nm) relates to their hydrodynamic radius (dH). For symmetrical particles in solutions DLS is perhaps the best method for measuring their size.
Characterising nanoscale systems
- Imaging/microscopy methods
- Optical microscopy
- Fluorescene microscopy
- Transmission electron microscopy (TEM)
- Scanning electron microscopy (SEM)
- Scanning tunneling microscopy (STM)
- Atomic force microscopy (AFM)
Characterising nanoscale systems
- Imaging/microscopy methods
- optical microscopy
Optical microscopy: Mild, easy to use. Doesn’t work on non-transparent samples. Resolution is > 200 nm – not overly useful for measuring sizes on the nanoscale.
Characterising nanoscale systems
- Imaging/microscopy methods
- fluorescence microscopy
Fluorescence microscopy: (including fluorescence confocal microscopy) – While suffering from the same resolution limits as optical microscopy, it is more useful in charactering systems on the nanoscale due to the sensitivity of fluorescence (see also two-colour methods below)
Characterising nanoscale systems
- Imaging/microscopy methods
- transmission electron microscopy
Transmission electron microscopy (TEM): Uses electrons rather than light for “illuminating’ the sample. Electrons that are transmitted through the sample are detected and form the image. Resolution < 1 nm (even 0.1-0.2 nm). Key tool for characterising nanoscale systems especially those that DLS is not got for, e.g. non-symmetrical ones, hollow particles if we want to measure membrane thickness or indeed any more complex nanostructures in chemistry and biology. Requires high vacuum. Contrast mainly comes from atomic composition – heavier elements give better contrast – most carbon-based (organic) material “look” similar. Therefore, we need to stain organic materials with heavy elements (W, U) but this can lead to artefacts. Samples has to be thin – no electrons will pass through (image will be “black”) if sample is much thicker than 100 nm.
Characterising nanoscale systems
- Imaging/microscopy methods
- scanning electron microscopy
Scanning electron microscopy (SEM): Related to TEM but measures electrons scattered, in all directions, from the sample. To create an image, the electron beam scanned (x, y- raster pattern) over the sample to generate the image. Great for surface structures. Sample in vacuum and often has to be coated with a very thin layer of gold, platinum or other heavy metal.
Characterising nanoscale systems
- Imaging/microscopy methods
- scanning tunneling microscopy
Scanning tunnelling microscopy (STM): The first scanning probe microscopy method invented. Atomic resolution (< 0.1 nm) is “relatively” straightforward. Samples can be measured in liquid. Only gives surface information. Requires conductive surface and/or sample.
Characterising nanoscale systems
- Imaging/microscopy methods
- atomic force microscopy
Atomic force microscopy (AFM): Much more versatile than STM; does not require a conductive surface. Measuring forces between a tip and surface. Can be used to image living cells or synthetic nanosystems. Only gives surface dimensions (height and x/y of some on a surface.
Characterising nanoscale systems
- Mass spectrometry methods
- Peptide sequencing mass spectrometry
- Two colour methods
- Co-localisation two colour
- Förster Resonance Energy Transfer (FRET)
Characterising nanoscale systems
- Mass spectrometry methods
- Peptide sequencing mass spectrometry
Proteins (which are nanoparticles!) can be sequenced with the aid of mass spectrometry by chopping up the protein into smaller peptide fragments and the perform mass spectrometry measurements that fragment these peptides further. The fragments are then puzzled together again to work out the sequence and hence mass of the protein.
Characterising nanoscale systems
- Mass spectrometry methods
- Two colour methods
Fluorescence microscopy is very good at telling us if two things (e.g. A and B) are near each or not. All we need to do is label these with different fluorescent dyes. And as long as both the spectral characteristics (excitation and emission) of the two dyes are different enough and the instrument configuration, especially the available emission bandgap filters, we can separate these two colours when we are imaging.
Some applications include: Seeing the difference between a nucleus and cytosol in a cell, seeing the difference between a living and a dead (dying) cell. Seeing if protein A is close to DNA B. Or if cancer drug A is bound inside polymer nanoparticle B.
Characterising nanoscale systems
- Mass spectrometry methods
- Two colour methods- Co-localisation two colour
Co-localisation two colour: This is the “conventional” approach. When selecting your dyes, you mainly have to ensure there is no cross-talk between the emission bands of these two dyes relative to the bandgap filters you use for fluorescence detection. N.b. if the two dyes appear together, it doesn’t mean that the two objects (A and B) are really close – they can easily be hundreds (Abbey diffraction limit), if not thousands of nanometers apart.
Characterising nanoscale systems
- Mass spectrometry methods
- Two colour methods- FRET
Förster Resonance Energy Transfer (FRET): When two dyes (fluorophores) are close enough (less than 10-20 nm), non-radiative direct energy transfer, so- called FRET, can take place between two dyes which are called the Donor (e.g. dye A) and Acceptor (e.g. dye B). This means exciting with wavelength specific to the Donor A can result in emission from B! The efficiency of FRET decays with distance – in fact it can be used to measure distances say within or between proteins.
When selectin dyes for FRET – similar
consideration regarding emission spectra and bandgap filter apply as in co-localisation method. In addition –you need to ensure the emission of the Donor matches the absorption/excitation of the acceptor!
