Choosing the Right Microscope

Compact electronic microscope

There are three main types of microscopes: optical, electronic and near field probe. These are their differences:

• Optical microscopes: they use optical lenses to form the image while controlling the light beam to light up the sample. Several parameters (the type of lighting, polarization, spectral filtering, spatial filtering) can be modified for different observation techniques. Digital microscopes are like a kind of optical microscope where the observer’s eye is replaced by a camera. Magnifications in optical microscopy are in the range of 1,000x.
• Electronic microscopes: the irradiation of the sample is carried out via an electron beam. The magnifications here are much larger than in optical microscopy and can reach up to two million times. There are two main types of electronic microscopes: transmission electron microscopes (TEM) and scanning electron microscopes (SEM).
• Near field probe microscopes:this type of microscopy consists of bringing a probe close to the surface of an object and determining the topography of the surface of a sample. Depending on the microscope used, the spatial resolution can reach the atomic level. Near field probe microscopes include, for example, atomic force microscopes (AFM) and scanning near-field optical microscopes (SNOM).

In the healthcare industry, microscopes are used in many fields such as:

• The medical field: microscopes are used for laboratory examinations, especially to study samples and detect the presence of pathogens such as microbes, bacteria or other potentially deadly micro-organisms. In histopathology, for example, microscopes are used to observe tissue samples for diagnostic purposes.
• Life sciences (biology): microscopes can be used to study microorganisms, for example their cellular make-up and development patterns.
• Forensic medicine: microscopes are used to analyze evidence found at crime scenes for the purpose of presenting evidence in court. Items such as dust, glass, bits of fabric, bodily fluids, hair, ink and microorganisms can be analyzed under a microscope.
• The pharmaceutical industry: microscopes are used in the pharmaceutical industry to ensure product safety and quality. Scientists analyze products under a microscope to detect any defects or contaminants.
• Teaching: many models are designed for use in practical exercises at schools and universities.
• Research 

There are many different possible observation techniques in microscopy, depending on the type of microscope used and the field of application. The most commonly used today are the following:

Confocal laser scanning microscopy

In optical microscopy:

• Bright field (conventional): is a conventional technique for collecting the white light transmitted by the sample. It does not require extensive sample preparation. On the other hand, it has low contrast and is unsuitable for thick samples.
• Dark field: here, the light emitted by the source is weakly transmitted to the microscope objective. Only the light scattered by the sample is collected by the microscope objective. It is intended to improve contrast and remove the need for staining samples. It remains unsuitable for thick samples.
• Fluorescence: light is emitted by the samples. These can be naturally fluorescent or made to be fluorescent. This is a very sensitive technique that is useful for marking and differentiating several structures simultaneously via their fluorescent colors. It is also effective for observing the biochemical dynamics of samples.
• Confocal laser scanning: the light source used is a laser. This technique is very often coupled with fluorescence microscopy. It offers better resolution, suitable for thick or spread out objects. It is also effective for fixed or dynamic observation (living cells for example). However, the equipment is very expensive and the laser used presents some risks.

Electronic microscopy:

Image of a blood clot taken by SEM technique

• Scanning (SEM): a fine beam of electrons is emitted on the sample to be observed. Their interaction leads to the formation of secondary electrons. These are detected and converted into signals that allow the relief image to be reconstructed with great depth of field. However, the preparation of the samples is onerous. They must be dehydrated and then undergo treatment to become conductive (tissue fixation, cleaning).
• Transmission (TEM): similarly, a fine electron beam is emitted onto the sample. However, in this case it’s the electrons that have passed through the sample that are detected by the microscope. They offer better resolution than miscroscopes using SEM. Samples must be prepared according to a specific protocol. They must retain their structure and be conductive to allow the electron beam to pass through.

In near field probe microscopy:

• Atomic Force Method (AFM): is a technique that uses the repulsive force between the electron clouds on the sample surface and the microscope probe’s electron cloud. This technique therefore allows the surface of a sample to be scanned, as the probe can move in all directions. The analysis of the path of the probe, as well as the measurement of the interaction forces between the probe and the sample, allows the surface topography to be defined. This method can be used for both conductive and non-conductive samples.
• Scanning near-field optical microscopy (SNOM): this is a super-resolution microscopy technique. It makes it possible to exceed the Abbe limit by detecting evanescent waves. The light detector is placed very close to the surface of the sample. This means that the evanescent wave is observed and not the scattered wave. Therefore, details smaller than the wavelength of light can be visualized.

Given the large number of applications for a microscope, the criteria vary greatly depending on the intended use. In general, you should consider five main characteristics when choosing a microscope: magnification, observation technique, configuration, number of eyepieces, and type of lighting.

• The total magnification of the microscope: this is the product of the magnification of the eyepieces and the objective.

• The microscopic observation technique

• The microscope configuration: most microscopes are upright, i.e. the light arrives from below the sample and the observation is done from above. There are also inverted microscopes where the light source is placed above the sample and the objectives are below.

• The number of eyepieces: there are monocular, binocular and trinocular microscopes.

• The type of lighting: most often the lighting is LED or halogen.

There are many compatible accessories available depending on the type of microscopy and the field of application concerned. They range from cameras to screens to zooms.

Here are some of the accessories available:

• Cameras: cameras are mounted on the objective and allow you to take photos of the samples being analyzed.
• Screens: they allow you to display what is observed under the microscope in real time for easy observation.
• Discussion bridge: this tool allows the beam to be split and displayed in two different objectives so that two users can simultaneously observe the sample.
• Slide scanners: these allow the observed sample to be scanned and the collected images to be processed by computer.
• Zoom: the zoom allows a continuous magnification of the observed sample.
• Motorization system: this is used to move the nose piece.