Phase contrast microscope

INTRODUCTION

It was first described in 1934 by Dutch physicist. Frits zernike.Phase contrast is light microscopy technique used to enhance the contrast of images of transparent and colourless specimen.Such as living cells (usually in culture) ,microorganism ,thin tissue slices, fibers ,subcellular particles including nuclei and other organelles.Phase contrast microscopy is an optical microscopy technique that converts phase shift in the light passing through transparent specimen of brightness changes in the image.

Principle

When light passes through cells , small phase shift occur ,which are invisible to human eye.In a phase shift microscope ,these phase shifts are converted into changes in amplitude,which can be observed as differences in image contrast.

Parts of phase contrast microscope

•Light source

•Annual diaphragm

•Condenser lens

•Specimen

•Objective lens

•Phase plate

•Ocular lens

It requires additional specialized structure annular diaphragm & phase plate.

•Annular diaphragm-It is situated below the condenser. It is made up of a circular disc having a circular annular groove . The light rays are allowed to pass through the annular groove . Through the annular groove of the annular diaphragm,the light rays fall on the specimen to be studied.

•Phase plate– The phase plate is also called as diffraction plate or phase retardation plate. It is located between the ocular lens & objective lens . The phase plate is a transparent glass disc with one or few channels. The channel is coated with a material that can absorb light but cannot retard it. The other portion (other than channel) of the phase plate is coated with light retarding materials such as Magnesium fluoride. Phase plate helps to reduce the phase of the incident light.

How does it works?

The light rays enter the annular diaphragm from its source. Then it passes through the condenser lens,which focused the rays on the specimen . As the light transmitted through the specimen it creates deviated and undeviated light rays. The light transmitted through the specimen and then enters the objective lens where an image of the specimen is created.The unretarded light rays are pass through the phase ring & the wavelength is enhanced 1/4. The retarded light rays are pass  through the phase  ring and the wavelength reduced by 1/4. The wavelength is changed so we can see clear image because total change in wavelength is  λ/2. These two rays are a joined together with high amplitude and past than through ocular lens which forms image.

Applications

•Determine morphologies of living cells such as plant & animal cells.

•Studying microbial motility and structures of Locomotion.

•To detect certain microbial elements such as the bacterial endospore.

•It is used in the diagnosis of tumor cells.

•Phase contrast enables visualization of internal cellular components.