Introduction Microscope

In biology, there are lots of things that are

interesting but too tiny to see well - sometimes

to see at all - with the naked eye. A

magnifying lens is good, but a series of lenses,

each magnifying the image of the last, works

better. That's a microscope.

Anything that you want to look at with a

microscope can be called your specimen, and

the nature of your specimen may dictate what

sort of microscope you need to see it with. If

you are looking at the outside, you want a

scope that scans the surface - a scanning

microscope images light (or other radiation)

reflected off the surface of a specimen. To

look inside, you need to get light (or other

radiation) to go through your specimen - it

needs to be thin, semi-transparent, or both. A

microscope that sees images passed through a

specimen is a transmission microscope.

History

The first microscope to be developed was the optical microscope, although

the original inventor is not easy to identify. Galileo is sometimes credited

with inventing the first simple microscope in 1610. Evidence points to the

first compound microscope appearing in the Netherlands in the 1620s,

probably an invention of eyeglass makers there. Two eyeglass makers there

are variously given credit: Hans Lippershey (who developed an early

telescope) and Zacharias Janssen (also claimed as the inventor of the

telescope). Robert Hooke is also cited as a possible inventor of the

compound microscope. There are other claims that the microscope and the

telescope was invented by Roger Bacon in the 1200s. Giovanni Faber coined

the name microscope for Galileo Galilei's compound microscope in 1625

(Galileo had called it the "occhiolino" or "little eye").

 

Types of microscopes

Electron microscopy

In the early 1900s a significant alternative to light microscopy was

developed, using electrons rather than light to generate the image. Ernst

Ruska started development of the first electron microscope in 1931 which

was the transmission electron microscope (TEM). The transmission electron

microscope works on the same principle as an optical microscope but uses

electrons in the place of light and electromagnets in the place of glass lenses.

Use of electrons instead of light allows a much higher resolution.

Light Microscopy

It was not until the 1660s and 1670s that the microscope was used

extensively for research in Italy, The Netherlands and England. Marcelo

Malpighi in Italy began the analysis of biological structures beginning with

the lungs. Robert Hooke's Micrographia had a huge impact, largely because

of its impressive illustrations. The greatest contribution came from Antonie

van Leeuwenhoek who discovered red blood cells and spermatozoa and

helped popularise microscopy as a technique. On 9 October 1676, Van

Leeuwenhoek reported the discovery of micro-organisms.[6]

In 1893 August Köhler developed a key technique for sample illumination,

Köhler illumination, which is central to modern light microscopy. This

method of sample illumination gives rise to extremely even lighting and

overcomes many limitations of older techniques of sample illumination.

Further developments in sample illumination came from Fritz Zernike in

1953 and George Nomarski 1955 for their development of phase contrast

and differential interference contrast illumination which allow imaging of

transparent samples.

 

Types of Light Microscopy

Bright field microscope

is the simplest of all the optical

microscopy illumination techniques. Sample illumination is transmitted (i.e.,

illuminated from below and observed from above) white light and contrast in

the sample is caused by absorbance of some of the transmitted light in dense

areas of the sample. Bright field microscopy is the simplest of a range of

techniques used for illumination of samples in light microscopes and its

simplicity makes it a popular technique. The typical appearance of a bright

field microscopy image is a dark sample on a bright background, hence the

name.

 

 

 

 

 

Dark field microscopy

(dark ground microscopy) describes microscopy methods, in both light and

electron microscopy, which exclude the unscattered beam from the image.

As a result, the field around the specimen (i.e. where there is no specimen to

scatter the beam) is generally dark.

 

 

 

Fluorescence Microscope

A fluorescence microscope is an optical microscope that uses fluorescence

and phosphorescence instead of, or in addition to, reflection and absorption

to study properties of organic or inorganic substances. The "fluorescence

microscope" refers to any microscope that uses fluorescence to generate an

image, whether it is a more simple set up like an epifluorescence

microscope, or a more complicated design such as a confocal microscope,

which uses optical sectioning to get better resolution of

the fluorescent image.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Phase contrast microscopy 

 

Phase contrast microscopy is an optical microscopy technique that

converts phase shifts in light passing through a transparent specimen to

brightness changes in the image. Phase shifts themselves are invisible, but

become visible when shown as brightness variations.

 

Figure 1: The same cells imaged with traditional bright field microscopy

(left) and with phase contrast microscopy (right).

When light waves travels through a medium other than vacuum, interaction

with the medium causes the wave amplitude and phase to change in a

manner dependent on properties of the medium. Changes in amplitude

(brightness) arise from the scattering and absorption of light, which is often

wavelength dependent and may give rise to colors. Photographic equipment

and the human eye are only sensitive to amplitude variations. Without

special arrangements, phase changes are therefore invisible. Yet, phase

changes often carry important information.

 

 

 

Polarised Microscopy

Polarised light microscopy uses plane-polarised light to analyse structures

that are birefringent; structures that have two different refractive indices at

right angles to one another (e.g. cellulose microfibrils). Normal, un-

polarised, light can be thought of as many sine waves, each oscillating at any

one of an infinite number of orientations (planes) around the central axis.

Plane-polarised light, produced by a polar, only oscillates in one plane

because the polar only transmits light in that plane.

Principles

The magnification of small things is a necessary facet of biological research,

but the fine detail in cells and in subcellular components requires that any

imaging system be capable of providing spatial information across small

distances. Resolution is defined as the ability to distinguish two very small

and closely-spaced objects as separate entities. Resolution is best when the

distance separating the two tiny objects is small. Resolution is determined by

certain physical parameters that include the wavelength of light, and the

light-gathering power of the objective and condenser

lenses. A simple mathematical equation defines the smallest distance (dmin)

separating the two very small objects:

dmin = 1.22 x wavelength / N.A. objective + N.A. condenser

This is the theoretical resolving power of a light microscope. In practice,

specimen quality usually limits dmin to something greater than its theoretical

lower limit. N.A. (Numerical Aperture) is a mathematical calculation of the

light-gathering capabilities of a lens.

 

 

References

1.     Microbiology - Krishna Prakashan Media, page 13

2.     Albert Van Helden, Sven Dupré, Rob Van Gent, Huib Zuidervaart,

The Origins of the Telescope, pages 32-36

3.     Microbiology - Krishna Prakashan Media, page 13

4. William Godwin (1876). "Lives of the Necromancers".

5. Gould, Stephen Jay (2000). "Chapter 2: The Sharp-Eyed Lynx,

Outfoxed by Nature". The Lying Stones of Marrakech: Penultimate

Reflections in Natural History. New York, N.Y: Harmony. ISBN 0-

224-05044-3.

6. Wootton, David (2006). Bad medicine: doctors doing harm since

Hippocrates. Oxford [Oxfordshire]: Oxford University Press. ISBN 0-

19-280355-7.

7. Knoll, Max (1935). "Aufladepotentiel und Sekundäremission

elektronenbestrahlter Körper". Zeitschrift für technische Physik 16:

467–475.

8. Morita S (2006). Roadmap of Scanning Probe Microscopy.

NanoScience and Technology. Berlin: Springer. ISBN 3-540-34314-8.

9. ^ Majumdar A (1999). "Scanning Thermal Microscopy". Annual

Review of Materials Science 29: 505–85.

Bibcode:1999AnRMS..29..505M.

doi:10.1146/annurev.matsci.29.1.505.

 

 

 

 

 

 

 

 

INDEX

 

       Introduction of Microscope

       History of Microscope

       Types of Microscope

1.   Electron Microscope

2.   Light Microscope

       Types of Light Microscope

1.   Bright Field Microscope

2.   Dark Field Microscope

3.   Fluorescence Microscope

4.   Phase Contrast Microscope

5.   Polarised Microscope

       Principle

       Reference

 

 

 

 

Acknowledgement

 

Preparing a project of this nature is an arduous task and I was

fortunate enough to get support from large number of person . I wish to

express my deep sense of gratitude to all those who generously helped in

successful completion of this report by sharing their invaluable time and

knowledge

It is my proud and privileged to express my deep regard to Respected Dr.

J.P.N . Pandey Principal of Govt. Girls P.G College of Excellence Sagar

Dr. Neera Sahay H.O.D. Zoology Department for allowing me to

undertake the project

I feel extremely exhilarated to have completed this project under the able

and inspiring guidance of Mr. Rakesh Kumar Saket he rendered me all

possible help and guidance while reviewing the manuscript in finalizing

this report .

I also extend my deep regards to my teachers , family members, friends and

all those whose encouragement has infused courage in me to complete the

work successfully.

 

Gulafsha

Kassab

B.Sc. IIIrd

Sem

 

 

 

Certificate

 

The project report titled “ Microscope ” in Sagar city prepared by

Gulafsha Kassab B.sc. (Biotech) IIIrd Sem. under the guidance and

supervision of Mr. Rakesh Kumar Saket , for partial fulfillment of the

Degree B Sc.

 

Signature of supervisor Signature of Examiner

………………… …………………

 

Signature of H.O.D.

…………………

 

 

 

 

 

 

 

 

 

 

 

Acknowledgement

 

Preparing a project of this nature is an arduous task and I was

fortunate enough to get support from large number of person . I wish to

express my deep sense of gratitude to all those who generously helped in

successful completion of this report by sharing their invaluable time and

knowledge

It is my proud and privileged to express my deep regard to Respected Dr.

J.P.N . Pandey Principal of Govt. Girls P.G College of Excellence Sagar

Dr. Neera Sahay H.O.D. Zoology Department for allowing me to

undertake the project

I feel extremely exhilarated to have completed this project under the able

and inspiring guidance of Mr. Rakesh Kumar Saket he rendered me all

possible help and guidance while reviewing the manuscript in finalizing

this report .

I also extend my deep regards to my teachers , family members, friends and

all those whose encouragement has infused courage in me to complete the

work successfully.

 

Sameeksha

Mishra

B.Sc. IIIrd

Sem

 

 

 

Certificate

 

The project report titled “ Microscope ” in Sagar city prepared

by Sameeksha Mishra B.sc. (Biotech) IIIrd Sem. under the guidance

and supervision of Mr. Rakesh Kumar Saket , for partial fulfillment

of the Degree B Sc.

 

Signature of supervisor Signature of Examiner

………………… …………………

 

Signature of H.O.D.

…………………

 

 

 

 

 

 

 

 

 

 

References

 1.      Southern, Edwin Mellor (5 November 1975). "Detection of specific

sequences among DNA fragments separated by gel electrophoresis".

Journal of Molecular Biology 98 (3): 503–517. doi:10.1016/S0022-

2836(75)80083-0. ISSN 0022-2836. PMID 1195397.

2.     Towbin et al.; Staehelin, T; Gordon, J (1979). "Electrophoretic transfer

of proteins from polyacrylamide gels to nitrocellulose sheets: procedure

and some applications". PNAS 76 (9): 4350. doi:10.1073/pnas.76.9.4350.

PMID 388439.

3.      Burnette, W. Neal (April 1981). "Western Blotting: Electrophoretic

Transfer of Proteins from Sodium Dodecyl Sulfate-Polyacrylamide Gels

to Unmodified Nitrocellulose and Radiographic Detection with Antibody

and Radioiodinated Protein A". Analytical Biochemistry 112 (2): 195–

203. doi:10.1016/0003-2697(81)90281-5. ISSN 0003-2697.

PMID 6266278.

4.      Biochemistry 3rd Edition, Matthews, Van Holde et al, Addison Wesley

Publishing, pg 977