- Introduction
A compound light microscope represents two or more lens. The term compound refers to multiple or mixture. In compound light microscope built with binocular lens and objective lens. The binocular lens present near the eyepiece and objective leans near to nosepiece. The objective lens helps in enlargement of images and also to collect the light source. According to the requirement the objective lens also changes. In advanced compound light microscope the phase contrasts also possible. In earlier 1595 Zacharias Janssen invented the first light microscope using collapsing tubes. It produced a magnification of 9x. Now the magnification power reached a long way i.e. nearly 2000x. The compound light microscope also named as the bright field microscope. The most important property of a microscope is its resolution capacity. These microscopes mainly used for study of plant cell, animal cell, bacteria, and protozoa. The compound light microscope classified in to two types they are upright microscope and inverted microscope. In upright microscope the parts are lens arrangement, specimen, and light source. In inverted the light source in the top then followed by specimen and lens. By changing the power of the lens not only the high resolution achieved we also increase the amount of specimen. The reflective index, angular aperture, and wavelength help in better result of compound light microscope. The compound light and electronic microscope varies with the structural designs and the construction but they both operate on the same principle of magnifying the specimen. The compound light microscope also includes the light source, a mirror, a nosepiece, adjustable knob, and an arm. By discussing the compound light and the electronic microscope the advantages and disadvantages of compound light microscope will be known. The parts description also helps in better understanding of the compound light microscope and lead to future innovation.
A simple arrangement of compound light microscope:
HUMANEYE OCULAR LENS INTERMEDIATE LENS OBJECTIVE LENS SPECIMEN CONDESNSER LENS LIGHT SOURCE.
A simple electronic microscope:
HUMANEYE PROJECTOR LENS INTERMEDIATE LENS OBJECTIVE LENS SPECIMEN ANODE CONDENSER LENS TUNGSTEN FILAMENT.
- Parts of compound light microscope:
The architecture that includes body tube, resolving nosepiece, objective lens, clips, diaphragm, illuminator, eyepiece lens, the arm, the stage, coarse focus, fine focus, and the base. The eyepiece lens is ocular lens it helps in view of specimen. According to our need the eyepiece lens varied. The body tube help by supporting mechanically to objective lens and the eyepiece. The body tube length is a fixed one and also the objective lens and eyepiece lens are aliened. Resolving nosepiece plays an important role in the process because according to the requirement it will change. It present near the objectives. The arm connects the base and top. Arm is a curved part. For good handling one hand supports on arm and other on base. The name objective refer that it is present near the object the objective lens present near the specimen and having a high magnifying capacity. The diaphragm is the important in the compound light microscope because the help to adjust the light intensity and also able to change according to the size of the specimen and it is a movable part, focus is its main duty. The coarse and fine are the two adjustment types in the microscope to adjust the stage and body tube. The fine is for fine focusing and precise of the object. The condenser lens collects the light and help for focusing it gives the clarity to the specimen. The base part consists of light source and the base supports the whole weight.
Compound light microscope are limited in the application but it also a powerful instrument. An oil immersion lens in light microscope helps in high magnification to view the bacteria. Due to its high magnification sometimes the specimen are thick viewed with many layers, So that the vision is inaccurate. For the high resolution electron microscope are used. An optimal light microscope gives better resolution today. A fluorescence light microscope is a type of light microscope with some advancement in the light microscope these microscopes use higher light intensity with a longer wavelength. This fluorescence creates a higher clarity. These are not mostly available for common peoples.
- Fluorescence microscope:
The fluorescence microscope employs light to excite fluorescence in the specimen. A fluorescence microscope has an exciter filter between the light source and condenser lens that transmits only light of particular wavelength. The condenser fluorescence compounds in the specimen to emit light of longer wavelength. Both the excitation light from the illuminator and the emitted light generated by fluorescence compounds in the specimen then pass through the objective lens. As the light passes through the tube of the microscope above the objective lens, it encounters a barrier filter that specifically removes the excitation wavelengths. This leaves only the emission wavelength to form the final fluorescence images, which therefore appears bright against a dark background.
The light microscope and electronic microscope have both strengths and weaknesses. In electron microscope, resolution is much better, but specimen preparation and instrument operations are often much difficult. It based on two basic designs: the transmission electron microscope and the scanning electron microscope. Scanning and transmission electron microscope are similar in that each employs a beam of electrons to produce an image. However, the instrument uses quite difficult mechanisms to form the final images.
The focal length is the distance from the midline of a lens to the point at which parallel rays passing through the lens converge to a focus. The angular aperture of a lens is the half angle of the cone of light entering the objective lens of the microscope from the specimen. A low aperture lens is small angular aperture when the angular aperture is high more information can be transmitted a best angular aperture of a glass is 70 degree.
In compound light microscope the light rays from the source first pass through condenser lenses, which direct the light toward a specimen mounted on a glass slide and positioned on the stage of the microscope. The objective lens, located above the specimen is responsible for forming the primary image. The objective lens of different magnification mounted on rotating turret help to choose the specimens needed magnification. The ocular lens in the eyepiece enlarges the image to larger size. An intermediate lens is placed between the objective lens and ocular lens helps in further enlargement. The overall image is enlarged by objective lens, ocular lens, and intermediate lens. Compared to other microscope, the bright field microscope is inexpensive and simple to align and to design. However, the only specimens that can be seen directly by compound light microscope are those that possess color or have some other property that affects the amount of light that passes through. Many biological specimens lack these characteristics and must therefore be stained with dyes. The special microscopes have various advantages that make them especially well suited for visualizing specific types of specimens. These include phase contrast microscopy, differential interference contrast microscopy, florescent microscopy, and confocal microscopy
- Advantages and disadvantages:
The advantages of compound light microscope are particularly in medical science. Because of its high magnification the cut slice of tissues are examined and to detect the disease and diagnose it. The smaller magnification helps in examine the external structures. In this way they help in the study of their structure and functions. The smaller size is also a big advantage of compound light microscope and the transportation is easy. They cost is not high so it’s easily affordable. When compared to electron microscope utilize a larger area with high investments.
Usually the light microscope uses the white light the wavelength is some microns. These light microscope have the capacity to view above the wavelength of white light tends to invisible image. So for small specimens the compound lights not preferred.
The compound light can magnify for smaller images tend to expensive of the lens so that the other microscopes are preferred. The specimen with shorter wavelength of visible light will be difficult to magnify.
During the magnification it provides a 3 dimension image with a one layer of blurred image which make difficult to examine the specimen. Because of the lens focal length varies with the 2 dimensional images.
The specimens mostly reflects the electromagnetic radiations such waves are not inferred by the compound light because they accept only the visible lights which makes the specimen not clearly visible and the colorless images will appear. We can overcome this limitations tend to an invention of the new low cost and technical evolution in the light microscope. These microscopes changes according to the specimens needs. Both the compound light and electron microscope have drawbacks.
- Conclusion
The fluorescent microscopy resolution tends to be 250nm which is a superior one and the spacing between the two objects seen separately. For a standard fluorescent light microscope it is an impossible due to its overlapping light wavelength. Phase contrast microscope detects difference in reflective index and thickness. Differential contrast microscope utilizes a split light beam to detect phase difference; fluorescence microscope can detect the presence of specific molecules or ions within cells. Confocal microscope minimizes blurring by excluding out of focus light from an image. Digital video microscope can record enhanced time lapse images; optical method can be used to measure the movement and properties of proteins and other macromolecules. These are types of compound light microscope. In electron microscope the techniques involves scanning electron microscope revels the surface architecture of cells and organelles. Corrective microscope used to bridge the gap between light and electron microscope. Stereo electron microscope and x- ray diffraction are the latest technology to view the specimen in three dimensions.
References
1. Anderson, T., 1951. TECHNIQUES FOR THE PRESERVAATION OF THREE-DIMENSIONAL STRUCTURE IN PREPARING SPECIMENS FOR THE ELECTRON MICROSCOPE*. Transactions of the New York Academy of Sciences, 13(4 Series II), pp.130--134.
2. Burton, E. and Kohl, W., 1942. The electron microscope. 1st ed. New York: Reinhold Pub. Corp.
3. Egger, M. and Petran, M., 1967. New reflected-light microscope for viewing unstained brain and ganglion cells. Science, 157(3786), pp.305--307.
4. Grimstone, A., 1977. The electron microscope in biology. 1st ed. London: Edward Arnold.
5. Huxley, H., 1963. Electron microscope studies on the structure of natural and synthetic protein filaments from striated muscle. Journal of molecular biology, 7(3), pp.281--30.
6. Johnson, B., 1949. A Compound Reflecting Microscope of High Aperture for use in Ultraviolet Light. Journal of Scientific Instruments, 26(5), p.148.
7. Kellenberger, E., Ryter, A. and S'echaud, J., 1958. Electron microscope study of DNA-containing plasms II. Vegetative and mature phage DNA as compared with normal bacterial nucleoids in different physiological states. The Journal of biophysical and biochemical cytology, 4(6), pp.671--678.
8. Lewis, A., Isaacson, M., Harootunian, A. and Muray, A., 1984. Development of a 500 AA spatial resolution light microscope: I. light is efficiently transmitted through $lambda$/16 diameter apertures. Ultramicroscopy, 13(3), pp.227--231.
9. Mittemeijer, E., 2010. Fundamentals of materials science. 1st ed. Berlin: Springer.
10. Sivasankar, B., 2012. Instrumental methods of analysis. 1st ed. New Delhi: Oxford University Press.
11. Slayter, E. and Slayter, H., 1992. Light and electron microscopy. 1st ed. Cambridge [England]: Cambridge University Press.
12. Stump, D., 1922. AN APPLICATION OF POLARIZED LIGHT TO RESOLUTION WITH THE COMPOUND MICROSCOPE.*. Journal of the Royal Microscopical Society, 42(3), pp.264--266.
