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LIGHT

MICROSCOPE
By Group 2 :
Norsyamira Ismail 1316028
Shahrani Bin Shahiran
Afzan Fahmie
Si? Natasha Azman 1313554
Nur Syahidah Bin? Sani Dhillon
Outline
History of Microscope
Concept of the compound microscope
Calcula6on of angular magnica6on
Part of op6cal microscope and its func6on
Dierent Type of Microscopy
Advantages of light microscope
Disadvantages of light microscope

Case study
Deni?on - light microscope

From the Ancient Greek:


mikrs, "small" and skopen "to look" or "see"
An instrument used to see objects that are too
small for the naked eye.
A "light" microscope is one that relies on light
to produce the viewed image.
hRps://www.emaze.com/@AWWIFLIZ/Un?tled
Basic terms used

Magnica6on - the ability to view an object as
larger

Resolu6on - the ability to dis?nguish two
objects as separate en??es, rather than
seeing them blurred

Light microscope
Found in most schools, use compound lenses
to magnify objects which is govern by op?cal
principles.
Op?cal lens : concave (diverging) and convex
(converging)

Image from hRp://www.bbc.co.uk/bitesize/standard/physics/health_physics/light_and_sound/revision/2/


They use convex lenses because they need to
focus the light into the eye. Convex focuses in
while concave spreads the light out. So, they
use convex lenses.

Concave lenses are not normally used in


microscope except with combina?on with
other lenses.
Back to History
Zacharias Jansen created a compound
microscope with two lenses

1565 Magnica?ons up to 9x

Antony van Leeuwenhoek invented a simple (one-lens) microscope


Craded beRer lens and magnied up to 20x and achieved twice the
resolu?on of the best compound microscopes
First to see individual cells, including bacteria, protozoan,
muscle cells, and sperm.

1670
Achroma?c lens was developed for use in eyeglasses by Chester
Moore Hall
It is because light refracted when passing through the lenses
and altered what the image looked like

1729

Robert Hooke further rened


the compound microscope,
adding such features as a stage
to hold the specimen, an
illuminator, and ne focus controls
1800
Magnica?ons of 30x to 50x

More stable and smaller
Lens improvements from people around the world
August Kohler developed the parfocal lens which
allow the specimen to remain in focus when changing
objec?ves on a microscope.
18th -19th Ernst Leitz inven?ng a way to
centuries photograph the specimens.

Carl Zeiss and Ernst Abbe added the sub-stage condenser and
developed superior lenses that greatly reduced
chroma?c and spherical aberra?on
Late 19th
Improved resolu?on and higher magnica?on
centuries up to 2000x

Concept of Compound
Microscope
SHAHRANI B SAHIRAN
1314173
Light is rst emiRed by the light source and is
directed by the condenser lens on to the
specimen.
The light from the specimen then passes
through the objec?ve lens.
Then, light rays are passed to a projector lens,
which reverses their direc?on so that when
the image reaches the eye it will not appear
upside-down.
Not all microscope have a projector lens, so the
viewer may be seeing a reverse image.
In these cases, when the slide is moved, it will
appear to be moving in the opposite direc?on to
the viewer.
The light rays then travel to the ocular lens or
eyepiece. This is oden a 10X magnica?on lens,
meaning it magnies the magnied image an
addi?onal ten ?mes.
The image is then projected into the eye.
Compound microscope: A microscope that
consists of two microscopes in series
the rst serving as the ocular lens with a longer
focal length (the eyepiece) and
the second serving as the objec?ve lens with a
very short focal length (close to the object to be
viewed).
There are two kind of compound light
microscope.
Mainly they dier in the source of light they
use.
One uses a mirror to converge sunlight
While other uses directly an illuminator
Mirror Type Light Compound Microscope
Illuminator Type Light Compound Microscope
Part and Its Func?on
Eyepiece and Body tube

Eyepiece is the lens
through which the
viewed looks to see
the specimen.
It is usually contains
a 10X or 15X power
lens.
The body tube
connects the
eyepiece to the
objec?ve lenses.
Objec?ves and stage clips
The objec?ve lenses
are the closest to the
specimen.
A standard
microscope has three
to four objec?ve
lenses which range
from 4X to 100X.
Stage clips are metal
clips that held the
slide in the place.
Objec?ves Lenses
LPO / Low Power
Objec?ve
Gives the lowest
magnica?on, usually
10X
HPO / High Power
Objec?ve
Gives higher
magnica?on usually
40X or 43X
OIO / Oil Immersion
Objec?ve
Gives the highest
magnica?on usually 97X or
100X and is used wet either
with cedar wood oil or
synthe?c oil.
Image of light compound microscope (from below)
Arm and base

The arm connects


the body tube to the
base of the
microscope.
The base supports
the microscope and
its where illuminator
located.
Illuminator and stage

Illuminator is the light
source for a
microscope
A compound light
microscope uses a low
voltage bulb as an
illuminator.
Stage is the at
plalorm where the
slide is placed.
Nosepiece and aperture

Nosepiece is a rota?ng
turret that holds the
objec?ve lenses.
The viewer spins the
nosepiece to select
dierent objec?ve
lenses.
The aperture is the
middle of the stage
that allows light form
the illuminator to
reach the specimen.
Condenser and diaphragm.
A condenser gathers and
focuses light from the
illuminator onto the
specimen being viewed.
Diaphragm is a ve holed
disk placed under the
stage.
Each hole is of a dierent
diameter. By turning it,
you can vary the amount
of light passing through
the stage opening.
Calcula?on of Magnica?on
Formula
Total magnica?on
Linear Magnica?on

Cont.

Type of microscopy
Dark eld and bright eld
Phase contrast
Polarizing
Fluorescent
Bright eld and dark eld
Bright eld Dark eld
The image appear The image appear
darker bright
The background is The background is
bright dark
To observe the To observe aqua?c
natural colors of organism: algae,
sample or planktons, bacteria
observa?on of
stained sample
Phase contrast
a special type of light
microscope that
increases dierences in
light and dark areas
enhances contrast of
transparent and
colourless specimens
primarily used in
biological and medical
research Stages of Cell
Division
Polarizing
Involving polarized light
Directly transmiRed
light can op?onally be
blocked with a polarizer
orientated at 90
degrees to the
illumina?on.


Fluorescence
A uorescence microscopy is an
op?cal microscopy that uses
uorescence and
phosphorescence to generate
image
specimens are self-illuminated by
internal light
bright objects are seen in vivid
color against a dark background
Structure and image of light microscope and electron
microscope
Work ow of light microscope and electron microscope
Comparison of light microscope and electron microscope

LIGHT MICROSCOPE ELECTRON MICROSCOPE


Advantage Disadvantage
cheap to purchase Expensive to purchase
Cheap to operate Expensive to operate
Small and portable Very large and must be operated in
special rooms
Prepara?on of material is rela?vely quick Prepara?on of material is lengthy and
and simple requires considerable exper?se
Material rarely distorted by prepara?on Prepara?on of material may distort it
Living as well as dead material may be Living material cannot be observed
viewed
Natural colour of material can be All image are in black and white
observed
Comparison of light microscope and electron microscope

LIGHT MICROSCOPE ELECTRON MICROSCOPE


Disadvantage advantage
Magnies object up to 1500X Magnies objects over 500000X
Can resolve objects up to 200 nm apart Has a resolving power for biological of
specimens of around 1 nm
The depth of eld is restricted It is possible to inves?gate a greater
depth of eld
Problem Statement
Assigned to do a group project for assignment
(SPS 4220)

Discussion on
Introduc?on of instrument
What type of sample we want to use?
Why we choose the sample?
Advantages compare to other
instrument;
Other op?on- need to change the
method, etc.
Objec?ves
1. Demonstrate the proper procedures used in correctly using the
compound light microscope
2. Prepare and use a wet mount
3. Determine the total magnica?on of the microscope
4. Explain how to properly handle the microscope
5. Describe the changes in the eld of view and available light when
going from low to high power using the compound light
microscope
6. Explain why objects must be centers in the eld of view before
going from low to high power using the compound light
microscope
7. Explain how to increase the amount of light when going from low
to high power using the compound light microscope
8. Explain the proper procedure for focusing under low and high
power using the compound light microscope
Case study 1:
Tap Water and Fresh Water Pond
Under Light Microscope

Aim:
To the study of content in tap water and pond
water under light compound microscope:

Tap water
Objec?ve lens: 4x
Eyepiece lens: 10x
Total magnica?on: 40x

Dark eld Bright eld


Tap water
Objec?ve lens: 10x
Eyepiece lens: 10x
Total magnica?on: 100x

Dark eld Bright eld


Tap water
Objec?ve lens: 40x
Eyepiece lens: 10x
Total magnica?on: 400x

Dark eld Bright eld


Pond water
Objec?ve lens: 4x
Eyepiece lens: 10x
Total magnica?on: 40x

Fresh pond water Preserved pond water (stain)


Pond water
Objec?ve lens: 10x
Eyepiece lens: 10x
Total magnica?on: 100x

Fresh pond water Preserved pond water (stain)


Observa?on
Tap water Pond water

1) At 40x, bubble appear because the 1) At 40x, unicellular organisms can be


coverslip is not in 45 degree. seen but cannot be iden?ed.
2) tap water content is basically 2) At 100x magnica?on, enables to
contaminant. observe the unicellular organisms
3) Maybe from carbon lter or iron due from Pro?sta (Kingdom); Ciliates
to construc?on, lead pipes corrode. (class: Ciliata) that is the paramecium.
Case Study 2:
Structure of Plant Cell and Animal Cell
Under Light Microscope
Type of microscope : Lieca model DM750
Specimen : Onion strip (bright eld microscopy)

Part and func?on of plant cell organelle

Type of microscope : Nikon Inverted Light Microscope
Specimen : Mouse kidney
Type of stain :Haematoxylin and Eosin stain (H&E stain).
Source: School of Anatomy and Human Biology - The University of Western Australia

Adjust the course knob until


Select the appropriate slide the specimen is in focus

Position specimen at center


Gently place the slide on the slide of field of view
stage and secure with slide clip

Adjust the fine focus knob


until the specimen is clear
Position the 4x objective lens in
place (low power lens)
Change to higher power
objective lens

Adjust the light control Repeat process at new


magnification level
Follow steps to focus using low power
Click the lens to the longest objec?ve
Do NOT use the Coarse Focusing Knob
Thats the big one
Use the Fine Focus Knob un?l you can see
specimen clearly
Thats the small one
Always carry with 2 hands
Only use lens paper for cleaning
Do not force knobs
Always store covered
Keep objects clear of desk and cords
Making a Wet Mount slide
1. Gather a thin slice/piece of your
specimen. If your specimen is too thick,
then the coverslip will wobble on top of
the sample like a see-saw:

Making a Wet Mount slide
2. Place ONE drop of water directly over
the specimen.

Making a Wet Mount slide
3. Place the coverslip at a 45 degree
angle (approximately), with one edge
touching the water drop, and let go.
Making a Wet Mount slide
Dropping the cover slip directly onto the
slide will cause air bubbles to be trapped
Conclusion &
Recommenda?on
1. Electron microscope against light microscope

2. Improvise and economize microscope (Glass bead microscope)