An ultramicroscope is a microscope with a system that lights the object in a way that allows viewing of tiny particles via light scattering, and not light reflection or absorption. When the diameter of a particle is below or near the wavelength of visible light (around 500 nanometers), the particle cannot be seen in a light microscope with the usual methods of illumination. The ultra- in ultramicroscope refers to the ability to see objects whose diameter is shorter than the wavelength of visible light, on the model of the ultra- in ultraviolet .
In the system, the particles to be observed are dispersed in a liquid or gas colloid (or less often in a coarser suspension). The colloid is placed in a light-absorbing, dark enclosure, and illuminated with a convergent beam of intense light entering from one side. Light hitting the colloid particles will be scattered. In discussions about light scattering, the converging beam is called a "Tyndall cone". The scene is viewed through an ordinary microscope placed at right angles to the direction of the lightbeam. Under the microscope, the individual particles will appear as small fuzzy spots of light moving irregularly. The spots are inherently fuzzy because light scattering produces fuzzier images than light reflection. The particles are in Brownian motion in most kinds of liquid and gas colloids, which causes the movement of the spots. The ultramicroscope system can also be used to observe tiny nontransparent particles dispersed in a transparent solid or gel.
Ultramicroscopes have been used for general observation of aerosols and colloids, in studying Brownian motion, in observing ionization tracks in cloud chambers, and in studying biological ultrastructure.
In 1902, the ultramicroscope was developed by Richard Adolf Zsigmondy (1865–1929) and Henry Siedentopf (1872–1940), working for Carl Zeiss AG. [1] Applying bright sunlight for illumination they were able to determine the size of 4 nm small nanoparticles in cranberry glass. Zsigmondy further improved the ultramicroscope and presented the immersion ultramicroscope in 1912, allowing the observation of suspended nanoparticles in defined fluidic volumes. [2] [3] In 1925, he was awarded the Nobel Prize in Chemistry for his research on colloids and the ultramicroscope.
Later the development of electron microscopes provided additional ways to see objects too small for light microscopy.
A colloid is a mixture in which one substance consisting of microscopically dispersed insoluble particles is suspended throughout another substance. Some definitions specify that the particles must be dispersed in a liquid, while others extend the definition to include substances like aerosols and gels. The term colloidal suspension refers unambiguously to the overall mixture. A colloid has a dispersed phase and a continuous phase. The dispersed phase particles have a diameter of approximately 1 nanometre to 1 micrometre.
Microscopy is the technical field of using microscopes to view objects and areas of objects that cannot be seen with the naked eye. There are three well-known branches of microscopy: optical, electron, and scanning probe microscopy, along with the emerging field of X-ray microscopy.
The optical microscope, also referred to as a light microscope, is a type of microscope that commonly uses visible light and a system of lenses to generate magnified images of small objects. Optical microscopes are the oldest design of microscope and were possibly invented in their present compound form in the 17th century. Basic optical microscopes can be very simple, although many complex designs aim to improve resolution and sample contrast.
In optics, any optical instrument or system – a microscope, telescope, or camera – has a principal limit to its resolution due to the physics of diffraction. An optical instrument is said to be diffraction-limited if it has reached this limit of resolution performance. Other factors may affect an optical system's performance, such as lens imperfections or aberrations, but these are caused by errors in the manufacture or calculation of a lens, whereas the diffraction limit is the maximum resolution possible for a theoretically perfect, or ideal, optical system.
Richard Adolf Zsigmondy was an Austrian-born chemist. He was known for his research in colloids, for which he was awarded the Nobel Prize in chemistry in 1925, as well as for co-inventing the slit-ultramicroscope, and different membrane filters. The crater Zsigmondy on the Moon is named in his honour.
A sol is a colloidal suspension made out of tiny solid particles in a continuous liquid medium. Sols are stable and exhibit the Tyndall effect, which is the scattering of light by the particles in the colloid. Examples include amongst others blood, pigmented ink, cell fluids, paint, antacids and mud.
The Tyndall effect is light scattering by particles in a colloid such as a very fine suspension. Also known as Tyndall scattering, it is similar to Rayleigh scattering, in that the intensity of the scattered light is inversely proportional to the fourth power of the wavelength, so blue light is scattered much more strongly than red light. An example in everyday life is the blue colour sometimes seen in the smoke emitted by motorcycles, in particular two-stroke machines where the burnt engine oil provides these particles. The same effect can also be observed with tobacco smoke whose fine particles also preferentially scatter blue light.
Nanoparticle tracking analysis (NTA) is a method for visualizing and analyzing particles in liquids that relates the rate of Brownian motion to particle size. The rate of movement is related only to the viscosity and temperature of the liquid; it is not influenced by particle density or refractive index. NTA allows the determination of a size distribution profile of small particles with a diameter of approximately 10–1000 nanometers (nm) in liquid suspension.
Dark-field microscopy describes microscopy methods, in both light and electron microscopy, which exclude the unscattered beam from the image. Consequently, the field around the specimen is generally dark.
Phase-contrast microscopy (PCM) is an optical microscopy technique that converts phase shifts in light passing through a transparent specimen to brightness changes in the image. Phase shifts themselves are invisible, but become visible when shown as brightness variations.
The optical properties of all liquid and solid materials change as a function of the wavelength of light used to measure them. This change as a function of wavelength is called the dispersion of the optical properties. The graph created by plotting the optical property of interest by the wavelength at which it is measured is called a dispersion curve.
Vertico spatially modulated illumination (Vertico-SMI) is the fastest light microscope for the 3D analysis of complete cells in the nanometer range. It is based on two technologies developed in 1996, SMI and SPDM. The effective optical resolution of this optical nanoscope has reached the vicinity of 5 nm in 2D and 40 nm in 3D, greatly surpassing the λ/2 resolution limit applying to standard microscopy using transmission or reflection of natural light according to the Abbe resolution limit That limit had been determined by Ernst Abbe in 1873 and governs the achievable resolution limit of microscopes using conventional techniques.
Time-lapse microscopy is time-lapse photography applied to microscopy. Microscope image sequences are recorded and then viewed at a greater speed to give an accelerated view of the microscopic process.
Super-resolution microscopy is a series of techniques in optical microscopy that allow such images to have resolutions higher than those imposed by the diffraction limit, which is due to the diffraction of light. Super-resolution imaging techniques rely on the near-field or on the far-field. Among techniques that rely on the latter are those that improve the resolution only modestly beyond the diffraction-limit, such as confocal microscopy with closed pinhole or aided by computational methods such as deconvolution or detector-based pixel reassignment, the 4Pi microscope, and structured-illumination microscopy technologies such as SIM and SMI.
A condenser is an optical lens which renders a divergent light beam from a point light source into a parallel or converging beam to illuminate an object to be imaged.
Differential dynamic microscopy (DDM) is an optical technique that allows performing light scattering experiments by means of a simple optical microscope. DDM is suitable for typical soft materials such as for instance liquids or gels made of colloids, polymers and liquid crystals but also for biological materials like bacteria and cells.
Light sheet fluorescence microscopy (LSFM) is a fluorescence microscopy technique with an intermediate-to-high optical resolution, but good optical sectioning capabilities and high speed. In contrast to epifluorescence microscopy only a thin slice of the sample is illuminated perpendicularly to the direction of observation. For illumination, a laser light-sheet is used, i.e. a laser beam which is focused only in one direction. A second method uses a circular beam scanned in one direction to create the lightsheet. As only the actually observed section is illuminated, this method reduces the photodamage and stress induced on a living sample. Also the good optical sectioning capability reduces the background signal and thus creates images with higher contrast, comparable to confocal microscopy. Because light sheet fluorescence microscopy scans samples by using a plane of light instead of a point, it can acquire images at speeds 100 to 1,000 times faster than those offered by point-scanning methods.
Live-cell imaging is the study of living cells using time-lapse microscopy. It is used by scientists to obtain a better understanding of biological function through the study of cellular dynamics. Live-cell imaging was pioneered in the first decade of the 21st century. One of the first time-lapse microcinematographic films of cells ever made was made by Julius Ries, showing the fertilization and development of the sea urchin egg. Since then, several microscopy methods have been developed to study living cells in greater detail with less effort. A newer type of imaging using quantum dots have been used, as they are shown to be more stable. The development of holotomographic microscopy has disregarded phototoxicity and other staining-derived disadvantages by implementing digital staining based on cells’ refractive index.
Henry Friedrich Wilhelm Siedentopf was a German physicist and pioneer of microscopy.
NanoSight Ltd is a company that designs and manufactures instruments for the scientific analysis of nanoparticles that are between approximately ten nanometers (nm) and one micron (μm) in diameter. The company was founded in 2003 by Bob Carr and John Knowles to further develop a technique Bob Carr had invented to visualize nanoparticles suspended in liquid. The company has since developed the technique of Nanoparticle Tracking Analysis (NTA), and they produce a series of instruments to count, size and visualize nanoparticles in liquid suspension using this patented technology.