The document summarizes the principles and applications of the phase contrast microscope. It describes how Fritz Zernike first developed phase contrast microscopy in 1934 and won the Nobel Prize for it in 1953. It works by converting invisible phase changes caused by cell structures into visible brightness differences using an annular diaphragm and phase ring. This allows transparent specimens like living cells to be viewed without staining. Common applications include examining live cells, blood samples, and other biological specimens. The phase contrast microscope provides rich, detailed images while allowing organisms to remain unfixed and alive.

1. SUMIT
180040420016
M.Sc. (Evn.)
Department of
Environmental
science and Eng. ,
G.J.U.& S.T., Hisar.
Principles and application phase
contrast microsope
Presented by…
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2. Contents
 Introduction
 Phase contrast microscope
◘ Principle
◘ working
◘ Applications
◘ Advantages
◘ Disadvantages
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3. The Phase contrast microscope
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 Phase contrast microscopy first described in
1934 by Dutch physicist Fritz zernike, Whom
awarded by Nobel prize in physics in 1953.
A phase contrast microscope makes it possible
by utilizing two characteristics of light,
diffraction and interference, specimens based
on brightness differences.
It requires additional specialized structure
annular diaphragm and phase contrast ring.

4. 4 Sources: www.biologydiscussion
.com/microscope/phase contrast-

5. Principle of Phase Contrast Microscope
It based on the wavelength (nature) of light rays
and the fact that light rays can be in phase or
out of phase.
Different shade of grey are distinguished to our
eyes due to differences in amplitude of light
rays.
PCM converts invisible small phase changes
caused by the cell component in to visible
intensity changes.
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6. In a Phase contrast microscope, one set of
light rays comes directly from the light
sources.
The other set comes from light that is reflected
or diffracted from a particular structure in the
specimen.
The images differences in refractive index of
cellular structure. Light passes through thicker
parts of cell is held up relative to the light that
passes through thinner parts of cytoplasm.
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7.  How does it work ?
 Light that does not interact with the speciman
is collected by the objective passes through
the phase ring, and is regarded exactly ¼
wavelength.
The Phase shifted is not detectable by the eye
so the resulting image on the image plane in
the microscope appears as a normal bright
background.
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8. Light passing through one material & into
another material of slightly different refractive
index or thickness will undergo a charge in
phase.
This charge in are translated into variations in
brightness of the structures.
 Natural light vibrates in many directions but
polarized light only one direction.
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9. 9 Sources :
https://www.bwoptics.com/newsend2asp

10. Application of Phase Contrast microscope
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transparent specimens such as living cells
 Most commonly used to provide contrast of
or
small organisms.
 Useful in observing cells cultured in vitro during
mitosis.
 Phase contrast enables visualization of internal
cellular components.
 It’s used in examination of growth, dynamics, and
behaviour of a wide variety of living cells in cell
culture.

11.  It applied for equipment from the study of
the living biological specimens, medical
applications, study of live blood cells, and
other biological and science applications.
 It’s used in diagnosis of tumour cells.
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12. ADVANTAGES
Unstained specimens can be viewed and imaged .
Live cells or organisms can be imaged in their natural
form without fixing or killing them.
Produces images that are information-rich and
detailed.
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13. DISADVANTGES
 Not useful to see the natural colours of specimens
or if staining is involved.
Each objective has its separate phase contrast filter.
Accurate measurements of samples cannot be
acquired to technical. limitations where light cannot
penetrate beyond a particular sample thickness.
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14. 1) Biophysics
Author : Vasantha pattabhi ,
N. Gautham.
Edition : 2009 ( Second )
2) Basic Biophysics For Biologist
Author : M. Daniel
Edition : 2003
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