Light Microscope (Brightfield microscope)
This is the most basic optical Microscope used in microbiology laboratories which produces a dark image against a bright background.
Made up of two lenses, it is widely used to view plant and animal cell organelles including some parasites such as Paramecium after staining with basic stains.
Working Principle of Light Microscope
The key steps in its operation are as follows:
1. Illumination:
Light from a source, usually a halogen or LED bulb, passes through a condenser lens. The condenser focuses the light onto the specimen on the slide. The light then passes through the transparent or semi-transparent specimen.
Abbe condenser is the condenser that concentrates and controls the light that passes through the specimen prior to entering the objective.
Ernst Abbe invented the Abbe condenser in 1870, and it’s still the basis for most modern light microscope condenser designs.
2. Magnification:
Once light passes through the specimen, it moves through a series of lenses, typically the objective lens and the eyepiece lens:
– Objective lens: Positioned close to the specimen, this lens provides the primary magnification. The light is refracted, creating a magnified image.
During visualization, the objective lens remains parfocal which means, when the objective lens is changed, the image still remains in focus.
– Eyepiece lens (ocular): This lens further magnifies the image formed by the objective lens.
What is seen in the microscope as an enlarged clear image of the specimen is known as the virtual image.
The total magnification is the product of the magnification of the objective lens and the eyepiece lens.
The magnification is standard, i.e not too high nor too low, and therefore depending on the magnification power of the lenses, it will range between 40X and 1000X.
3. Resolution:
The light microscope has a resolving power, determined by the numerical aperture of the lenses and the wavelength of light used. It determines the microscope’s ability to distinguish between two close points as separate entities.
A higher numerical aperture means that the lens or optical system can gather more light, which generally results in better resolution and brightness.
4. Focusing:
The microscope uses focusing mechanisms, such as coarse and fine adjustment knobs, to move the stage or the objective lenses, allowing the image to be brought into sharp focus. This ensures clear observation of the specimen.
5. Image Formation:
The final image seen by the observer is inverted and magnified due to the combined effect of the objective and eyepiece lenses. The light microscope can also be equipped with different filters to enhance contrast, such as in phase contrast or fluorescence microscopy.
Applications of Light Microscope
Light microscopes are widely used in various scientific fields due to their ability to observe cells and tissues in detail. Here are some significant applications in microbiology and other disciplines:
1. Cell Observation:
Light microscopes are fundamental in microbiology for observing the morphology and structure of cells. They allow microbiologists to view bacteria, fungi, protozoa, and other microorganisms, making them essential for identifying different species based on their shapes and arrangements.
2. Staining Techniques:
Many microorganisms are difficult to see under a microscope due to their transparent nature. Various staining techniques like Gram staining, acid-fast staining, and spore staining can be used with light microscopes to enhance contrast, helping to differentiate and classify microorganisms.
3. Tissue Examination:
In histology and pathology, light microscopes are used to examine tissue sections. Thin slices of tissue are stained and examined to detect structural changes, cell abnormalities, and diseases.
4. Studying Cell Division:
Microbiologists use light microscopes to study mitosis and meiosis, observing the different stages of cell division in various organisms. This is particularly important in understanding growth and reproduction in microbes.
5. Microbial Cultures:
Microscopy is essential in studying microbial growth. Scientists can examine microbial colonies grown on agar plates and observe the detailed structure of microbial cultures under the microscope.
6. Environmental Microbiology:
In environmental microbiology, light microscopes are used to examine soil, water, and air samples for microorganisms. They allow researchers to observe microbial communities, measure cell counts, and study the role of microorganisms in ecosystems.
7. Medical Diagnostics:
In clinical laboratories, light microscopes are used to identify pathogenic microorganisms in patient samples. Blood smears, urine, sputum, and tissue biopsies can be examined to detect infections or other medical conditions.
8. Education and Research:
Light microscopes are widely used in educational settings to teach students about the basics of microbiology and cell biology. They also play a crucial role in ongoing research by allowing scientists to observe microscopic structures and study their functions.
9. Plant and Animal Cells Study:
In botany and zoology, light microscopes are used to study the anatomy of plant cells (such as cell walls, chloroplasts, and vacuoles) and animal cells (such as cell membranes, nuclei, and mitochondria).
10. Microbial Motility:
Microbiologists can use light microscopes to observe motility in microorganisms, such as flagellar or ciliary movements, which is crucial in understanding their behaviour and interaction with their environment.
