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WHAT IS CONFOCAL MICROSCOPY?
Ans: -The concept of confocal microscopy toas initially doped by Marvin Minsky in the 1950s, at Harvard University with an aimelving the neurul network without staining the tissues but it did not bort due to lack of enough light source and a computerized system to store the large data.The work was later adapted by David Egger M, and dojmir Petran, forming a multiple-beam confocal microscope in the late 1960s they used a spinning disk known as Nipkow which they used to examine brain tissues and ganglion cells that were unstained. The technique was late mofed and published by Eggert forming a mechanical scanned confocal microscope, that was able to visualize images of cells. Later developments in science eluding the development of computers and laser technology and digital manipulons of images using algorithms, grew the advances in confocal microscopy, joten practically usable confocal microscopes by a series of scientists including red Brakenhoff (1979), Colin Sheppard, Tony Wilson, Brad Amos, and form White (1980s).
The first commercial focal microscope was developed in 1987 with improved optics and electronics, powerful lasers with high scanning efficiency. The modern confocal Microscope has all the possible integration of technology and mechanical components including optical components, which perform the primary function of the configuration by use of electronic detectors, a computer, and laser systems.
The functioning of the confocal microscope is a collective role of all its components to price an electronic image. To date, these microscopes have been used to investigate molecules, microbial cells, and tissues.
PRINCIPLE OF THE CONFOCAL MICROSCOPE :-
Principle of the Confocal Microscope Normally a conventional (wide field) Microscope different wavelengths from a light source, to visualise and illumina farge area of a specimen, forming fuzzy, murky, and crowded images bene cell sample images are captured from all directions, without a focal point
To avoid these issues, a Confocal ice is used in wide field or fluorescent microscopes, the whole specimen receives light, receiving complete excitement and emitting light whigh is detected by a photodetector on the microscope. However, with the confocal microscope, point illumination is the principle working mechanism.
A specimen is stained with fluore rome is examined. When a beam of light is focused at a particular point of the fluoro-chromatic specimen, it produces an illumination that is focused by the objective lens to a plane above the objectives. The objective has an aperture on the focal plane located above it, which primarily functions to block any stray light from reaching the specimen. A measure of the illumination point is about 0.25 to 0.8 um in diameter, determined by the objective numerical aperture and 0.5 to 1.5 um deep, with the brightest intensity.
The specimen normally lies between the camera lens and the perfect point of focus, known as the plane of focus. Using the laser from the microscope, the laser scans over a plane on the specimen (beam scanning0 or by moving the stage (stage scanning). A detector then will measure the illumination producing an image of the optical section, scanning several optical sections, they are collected in a computerized system as data, forming a 3D image. The image can be measured and quantified.
Its outcome is also focored by the aperture found aboue the objective which →Images produced by the confocal microscope have a very good contruct and blocks strug light resolution capacity despite the thickness of the specimen Images are stored in the high-resolution 3D image of the cell complexes including structures
→The main characteristic of the Confocal Microscope is at detects what is focused and anything outside the focus point, appears b The image of the specimen is formed when the microscope Zanner, scans the focused beam across a selected area with the contr of two high speed oscillating mirrors. Their movement is facilitated by qulianometer motors. One mirror moves the beam from left to right on the uterol X-axis while the second mirror translates the beam along the Y-axis. After a can on the X-axis, the beam moves rapidly back to the starting point to a new scan, a process known as flyback No information is collected during the fuck process, therefore the point of focus, which in the area of interest is what illuminated by the laser scanner.