Microscopy is the science of viewing objects that are very minute and can only be viewed by an instrument called a microscope. The science began in the 16th century and has been progressing into viewing specimens at higher resolutions, greater magnifications, and in 3-D. These advancements are in the three sub-branches of microscopy which are scanning-probe, optical, and electron microscopy. The paper analyses the latest advancements in the three fields indicating what the invented microscopes can be used in science.
In Biology, Microscopy is the technique of using microscopes to view objects that are very minute and can only be clearly viewed by an instrument called a microscope (Mertz, 2009). The three major branches in this field are scanning-probe, light, and electron microscopy that involve the usage of microscopes that have the same name as the sub-fields. The history of the microscope—instrument in microscopy—can be traced in 1590 at Middelberg, Netherlands when two scientists -- Zacharias Jansen and his father Hans—experimented with lenses in a tube that made objects appear larger (Mertz, 2009). The microscope they made was referred to as a compound microscope and it proved a foundation to Galieleo who furthered the research by adding a focusing device in the apparatus.
However, for several years little advancements were done on the microscope in the late 19th century when August Kohler came up with a method for illuminating samples hence the basis of optical microscopy. Further developments in light microscopy were achieved in the 1950’s when two contrasting techniques were developed, that are, differential and phase contrasts (Mertz, 2009). In the field of electron microscopy, the technology began in the early 20th century where electrons replaced light to view images. The technology came into prominence after World War II with the gadget being commercially used in 1965 as the ‘Sterioscan.’ Other latest advancements came in the late eighties with the atomic microscope and the scanning-probe microscope.
This paper will highlight the latest advancements in microscopy through describing some of the latest microscopes and techniques. The discussion will be discussed based on the sub-fields of microscopy that are based on technology used in imaging. A further analysis of modern improvements is also done with a comment on what types of investigations that can be done by the new microscopes.
The science involves directing light reflected from an object or light passing through an object through lenses of a microscope resulting in magnification of the object. Images in the past were viewed on a photographic plate but in recent advancements the image is captured digitally. This has resulted in the digital microscope that uses CCD (CHARGE COUPLED DEVICE) camera that enhances focusing of images (Mertz, 2009).
Digital microscopes are currently used to create a 3-D focus on objects which has been a welcome advancement in light microscopy that was seen to be shallow in terms of focus. The technology was developed at the University of Illinois and uses an imaging technique that creates three dimensional views of objects updating them several times per second (Mertz, 2009). This is considered an advancement in optical microscopy as the technology could be used in biological research where, for instance, embryologists could use the technique to analyse cells in growth of internal organs of an embryo.
Confocal-microscopy technique a new advancement, has also been used in focusing 3-D objects providing a way of viewing objects very deep in living and fixed tissues and cells (Mertz, 2009). Furthermore, the technology affords the capability of obtaining precisely distinct optical sections from which three dimensional images can be created. Principles and applications of confocal microscopy have been increasingly being used by researchers requiring single-laboratory microscopes. Advancement of microscopes using this technology are on the rise majorly because of new and advanced trends in ICT, laser systems, interference filters, detectors, and in highly specific targets consisting of fluorophores (Merz, 2009).
In the sub-field of light microscopy there reached a time in the mid-seventies when the limit for resolution in optical microscopy was obtained. This however was not taken as an obstacle but rather it paved the way for research in sub-diffraction microscopy techniques that have resulted in microscopes using the techniques presently. The diffraction barrier is broken using techniques such as (Mertz, 2009): STED (STIMULATION EMISSION DEPLETION); Near-field scanning; optical nano-antennas; PSF (POINT SPREAD FUNCTION); and PALM (PHOTO-ACTIVATED LOCALIZATION TECHNOLOGY).
The electron microscope was invented to solve the problems that optical microscopes exhibited. The instrument works with a beam containing numerous electrons that would illuminate objects and magnify them. “Although expensive, they have vast resolving powers than optical microscopes since electrons have wavelengths 105 shorter than visible light (Spence, 2009, p. 6)”. Researchers using the electron microscope mostly use two types which are TEM (Transmission Electron Microscope) and SEM (Scanning Electron Microscope).
The original and commonly used electron microscope was the TEM which had a problem in dealing with spherical aberration (Spence, 2009). Latest advancements have seen the problem being corrected with correctors hence resulting in higher resolutions up to below 50 Picometers (Spence, 2009). These high resolutions are very applicable in physical and biological sciences such as visualizing the structure of compounds, pathogens, viruses, and even atoms. Latest Modifications in TEM involve the use of cryogen hence cryomicroscopy that is essential in maintaining specimens at nitrogen/helium temperatures. Therefore imaging specimens can also be prepared in sub-zero temperatures which is crucial in trying to research individual molecules or compounds.
In the SEM, there is greater imaging and focus since the beam of electrons is directed towards a rectangular area of an object (raster scanning). Although the resolution is poorer in SEM than in TEM, there is greater depth of field, and good 3-D representations (Spence, 2009). SEM is used in fields such as: medicine to analyse samples of healthy and unhealthy specimens; forensic to examine evidence; metal to compare strengths in different conditions; and in scientific research to compare different samples (Spence, 2009). In electron microscopy there has also been an advancement that involves the combination of SEM, and TEM coming up with a LVEM microscope (LOW-VOLTAGE ELECTRON MICROSCOPE). This technique reduces tissue damages as voltage used is less than 5kV and further increases image contrast that is very useful in biological researches (Spence, 2009).
Scanning Probe Microscopy
“Scanning-Probe Microscopy is the latest branch of microscopy where images of objects are obtained using a probe that is made of gold, platinum/iridium or Silicon Nitride tips (Meyer et al., 2004, p.7)”. The first to be invented was the Scanning Tunneling Microscope whose probe involved the technology of quantum tunnelling. However, the limit of specific specimens and lesser resolution led to the advancement of using AFT (Atomic Force Microscope).
The AFT is very crucial in calculating, representations, and controlling nanotechnology materials (Meyer et al., 2004). It uses piezzo (pressure) electric scanners that aid in tiny but accurate probing of objects to come up with higher resolution images. Latest advancements involve the use of current at the tip of the probe to come up with resolutions that match SEM and TEM electroscopes (Meyer et al., 2004). This advancement has enabled the microscope become useful in imaging live tissues; studying biological macromolecules; and even objects in liquid environments (Meyer et al., 2004). Other advancement in this sub-field is the use of Photonic force microscopy (PFM) that uses the technology of optical-tweezers instead of piezzo or quantum tunneling making it very sensitive to environmental conditions (Meyer et al., 2004). This technology enables the possibility of investigating and analysing environments such as agarose that can be filled by particles.
It can be noticed that microscopy advancements stagnated since its origin in the 16th century, but in the beginning of 20th century there was a revolution in the science leading to advancements in electron, optical, and probe scanning microscopy. It is now possible to view object in 3-D using the three microscopy techniques with very high resolutions that can view objects less than 50 picometers. “There is research still being carried out with sub-fields such as Ultra-sonic Force Microscopy, infra-red microscopy, and digital holography technology that would ensure better imaging techniques in the future (Spence, 2009, p. 102)”.
Mertz, J. (2009). Introduction to Optical Microscopy. New Jersey: Roberts. Pp. 3-78
Meyer, E., Hug, J., & Bennewitz, R. (2004). Scanning Probe Microscopy: the world on a tip. New York: Springer. Pp. 7-89
Spence, J. C. (2009). High Resolution Electron Microscopy (3rd ed.). New York: Oxford University Press. Pp. 6+