Phase contrast microscope

Phase contrast microscope, Types of phase contrast, Parts of Phase contrast microscope, Application of Phase contrast microscope

1. Phase contrast MICROSCOPE R. NITHYA M. Sc., M. Phil ., (Ph. D) ASSISTANT PROFESSOR IN BIOTECHNOLOGY, SRI ADI CHUNCAHNAGIRI WOMEN’S COLLEGE, CUMBUM, THENI DT, TAMIL NADU, INDIA.

2. PHASE CONTRAST MICROSCOPE The phase contrast microscope is a light microscope. It is modification of compound microscope. It contains all the component of a compound microscope in addition to an annular ring and a phase plate . It magnifies not only object but also changes in brightness .

3. PRINCIPLE The phase contrast microscope separates the illuminating background light and the specimen scattering light . Phase contrast microscope is used to visualize transparent, colourless , unstained, living biological specimens . These objects are called phase objects . Light is bend (diffracted) and retarded based on the refractive index of the object. Highly refractive structures bend and retard light much. This principle is used in phase contrast microscope.

4. Phase contrast microscope is the first microscopic method which allow the observation of living cell . It was invented by Frits Zernike and was awarded noble prize in 1953 .

5. TYPES OF PHASE CONTRAST MICROSCOPE There are two types of phase contrast microscope. They are Positive phase contrast or Dark contrast Negative phase contrast or light contrast Positive phase contrast or Dark contrast In positive phase contrast, the specimen is dark and the background is bright. The specimen is surrounded by a bright halo. Negative phase contrast or light contrast In negative phase contrast, the specimen is bright and the background is dark. The specimen is surrounded by a dark halo.

6. STRUCTURE The phase contrast microscope has the following parts . Light source – Tungsten lamp Annular diaphragm Condenser Objective lens with phase plate Eye piece Body tube Coarse adjustment Fine adjustment Stage Stage clips Nose piece Arm Inclination joint base

7. ANNULAR DIAPHRAGM The annular diaphragm is a plate having circular thin area in the centre surrounded by thick areas. Hence , light coming from the source can pass through the diaphragm in the form of a circle.  

8. ANNULAR PHASE PLATE The annular phase plate is a transparent disc with a groove. The size of the groove is identical to the circular area of the sub-stage diaphragm. The annular groove of phase plate filters off 70 – 90% S –waves and leaves D-waves as such. It is placed at the back focal plane of the objective.

9. CONDENSER It gathers and focuses the reflected light on the object. The condenser has a diaphragm . It allows the required intensity ( power, concentration, strength ) of light to pass through.

10. DIAPHRAGM Diaphragm is a five holed disk placed under the stage. Each hole is of a different diameter. By turning it, you can vary the amount of light passing through the stage opening.

11. NOSEPIECE AND APERTURE. • Nosepiece is a rotating turret that holds the Objective lenses. The viewer spins the Nosepiece to select different Objective lenses. The Aperture is the middle of the stage that allows light from the Illuminator to reach the specimen.

12. OBJECTIVE LENS Metal cylinders attached below the nosepiece and contains especially ground and polished lenses It magnifies the object . Onion cells

13. TYPES OF OBJECTIVE LENSES LPO / Low Power Objective Gives the lowest magnification, usually 10x HPO / High Power Objective Gives higher magnification usually 40x or 43x OIO / Oil Immersion Objective Gives the highest magnification, usually 97x or 100x, and is used cedar wood oil or synthetic oil for better resolution.

14. EYE PIECE It magnifies the image produced by the objective lens.

15. BODY TUBE It is a tube with the objective lens at the lower end and the eye piece lens at the upper end.

16. COARSE ADJUSTMENT It moves the body tube up and down rapidly to correct the distance from the object to get focusing. FINE ADJUSTMENT It moves the body tube up and down slowly to make exact focusing.

17. SPECIMEN STAGE It is a platform with a hole in the centre. The light falls on the object through the hole. The slide is placed on the stage.

18. STAGE CLIPS Stage clips hold the slide firmly on the stage

19. NOSE PIECE It is in the form of rotating disc having holes for fitting the objective lenses .

20. INCLINATION JOINT It permits tilting of upper part of the microscope to adjust to the eye level.

21. BASE OR FOOT The foot keeps the body in position.

22. USES OF PHASE CONTRAST MICROSCOPE. • It is used to view the cellular parts of Protozoa, bacteria and others with out staining. .

23. • Employed to observe cells in the culture without fixation and staining

24. Used to examine water and other fluids for the presence of minute transparent microbes ( bacteria and flagellates)

25. Used to study phagocytosis , pinocytosis , cell divisions etc in cells.

26. Used to study the behavior of chromosome during mitosis and meiosis

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