Microscope, as evident from its name, is a device that enables us to see objects that are invisible to the naked eye. The name literally roughly translates to seeing small objects, where Micro means ‘small’ and scope means ‘to see’ (derived from Ancient Greek).
Microscope is an instrument used in the field of basic and applied sciences to observe microscopic cells. The naked or unaided human eye can see the objects, which are more than 0.1 mm in size. The function of the microscope is to enlarge, magnify and capture the images of the microscopic cells. The efficiency of microscopes is indicated by its resolution. Resolution is ability of microscope to distinguish two close related points.
Antony Van Leeuwenhoek invented the first simple microscope in 1683. In simple microscope, single lens is used for magnification whereas in compound microscope, multiple lenses are used. Monocular microscope has one eyepiece and binocular has two eyepieces. As research has advanced, we now have different types of microscopes that are commercially available.
Principles of Microscope
The microscope works on three principles of physics:
- Resolving power
- Numerical aperture
- Magnification: It is the ability of lenses to enlarge an object visually. If the magnifying power of lens is 10X it means that the given lens can enlarge the object up to 10 times. In compound microscope, the magnification is the product of magnifying power of both the lenses. The magnifying power of lens depends on focal length. Lower the focal length, higher is the magnifying power of lens.
- Resolution: The resolving power is the ability to distinguish two closely placed points. The resolution power of lens allows us to observe the details of an object. The resolution of microscope can find out by using Abbe’s equation.
d – distance between two closely distant points
λ – wavelength of light
n sin θ – numerical aperture
The microscope with higher magnification has small d value. λ is the wavelength of light, shorter is the wavelength; higher is the resolution. The wavelength of visible light is from 300 to 700 nm. The best resolution for light microscope is obtained in the range of 450 to 500 nm. ‘n’ is the refractive index of medium. Refractive index is the ability of the medium to bend the light. The angle of cone of light is affected by the refractive index of medium. The refractive index of air is 1. ‘θ’ is the half of the angle of the cone of light that enters the microscope. The value of ‘Sin θ’ cannot be more than 1 because angle of entering cone of light cannot be more than 90° and value of sin 90 is 1.
d= 0.5 x 450 nm
d= 225 nm 0r 0.2 μm
Hence, the resolution limit of light microscope is 0.2 μm.
Parts of Microscope:
The body is made from metal and the parts are divided into mechanical and ocular parts.
- Base: The bottom part is called as base. It gives support to the body. The illuminating source of microscope is placed at the base of microscope
- Arm: It connects the ocular part with the base. The arm is used to hold and carry the microscope. The coarse and fine focusing knobs are placed on the arm.
- Eyepiece: It is present at the top of microscope. It is the part through which we observe the sample/object. The magnifying power of the eyepiece lens is generally 10X.
- Eyepiece tube or body tube: The function of eyepiece tube is to hold the eyepiece and hence it is names as eyepiece tube.
- Nosepiece: This part connects the eyepiece tube to objective lenses. The flexibility of nosepiece allows switching the objective lenses.
- Objective lens: In compound microscope, in general 3 objective lenses are placed. Their magnifying power is 10X, 40X and 100X respectively. The total magnification of respective lenses would be 10X x 10X = 100X, 40X x 10X = 400X and 100X x 10X = 1000X (the magnifying power of eyepiece lens is 10X).
- Focusing mechanism (Adjacent knobs): The coarse and fine adjustment knobs are used for right placing of sample and for focusing it.
- Stage: It is place where sample or object is kept for viewing. For accurate placement, the stage is provided with clips that hold the slide firmly. The adjustment knobs are used to move the clips as per requirement.
- Aperture: It is present on the stage that allows the entry of light from illuminator to fall on sample or object.
- Source of Illumination: In non-electric microscope, the sunlight is used as source of illumination and hence mirror is placed to focus the sunlight. In electric microscope, lamp is placed of specific wavelength.
- Condenser: It is placed at the bottom of the stage. Its function is to focus the light on sample form illuminator. The lens quality of condenser equally affects the superiority of image.
- Diaphragm: It is generally associated with condenser. And it is also placed at the bottom of the stage, beside condenser. The diaphragm and condenser together produce hollow cone of light that strikes the sample and illuminate it.