01/06/25

2.1.1 Microscopes
Do now (5 minutes):
1) What two things does a microscope do?
2) What is the stage used for?
3) What is a sub-cellular structure?
3)

Challenge: What is the function of ribosomes?

STOP
 Do
Do now corrections (in green pen)
now corrections
1) What two things does a microscope do?
Focuses and magnifies
2) What is the stage used for?
To hold the slide that will be viewed under the microscope
3) What is a sub-cellular structure?
A structure within a cell
2)

Challenge: What is the function of ribosomes? Makes proteins

 This lesson: Microscopes

•
• To understand the difference between
magnification and resolution
• To be able to describe different types of
microscopes
•

This half term

This Unit – Cell structure 2.1

2.1.1- Microscopes
2.1.2- Slides and photomicrographs
2.1.3- Measuring objects seen with a light
microscope (PA)
2.1.4- The ultrastructure of eukaryotic cells
PAG 1.1- Microscopy
2.1.5- Other features of eukaryotic cells
2.1.6/2.1.7 (PWFT)
2.1 Revision
Who is this and what happened to
him?
• In 1995 Christopher Reeves best known for playing
Superman, fell off his horse and had a spinal
injury. He became paralysed and wheelchair
bound
• He spent a lot of money on research into stem cells
• Ethical concerns were raised about embryonic stem
cells.
• In 2006 in Japan, a team found that they could
reprogram human skin cells to become stem
cells.
• As technology improves, we now are able to learn a
lot more about cells and further detail about
structures within cells
What you should know from GCSE’s
• Cells are the building blocks of life
• Metabolic processes like respiration are carried out in cells
• Cells are small so can only be seen with microscopes
• All cells have common features such as; surface membrane, DNA
• There are differences between plant, animal and bacterial cells
 Think, turn & talk task

How do we get these

images? What are
these images of?
How are they
different?

How big are these

things?
What is this?
What’s the difference between
magnification and resolution?
• Magnification – describes how much bigger an image appears
compared with the original object. Microscopes produce linear
magnification, which means that if a specimen is seen magnified
x100, it appears to be 100 times wider and 100 times longer than it
really is.
•
• Resolution – is the ability of an optical instrument to see or produce
an image that shows fine detail clearly. You may have a high-
resolution television (called ‘ultra-high definition’ or UHD) and have
noticed how clear and sharp the images on its screen are. Higher
resolution = clearer image.
•
Contrast
Contrast is difference in colour/shade between two objects
How to use a light microscope
1
2

6
Put the statements in order of
how to use a microscope
1. Make sure that the object you wish to view is directly over the hole in
the stage. Now rotate the nosepiece and bring the x10 objective into
place over the specimen. Look down the ocular tube and use the fine
focus knob to focus the image.
2. The specimen on a slide is placed here on the stage and clipped into
place.
3. Whilst viewing the image adjust the iris diaphragm for optimum light.
4. Repeat step 5 using the x40 objective lens.
5. By rotating the nosepiece, the lowest power (smallest) objective lens is
placed over the specimen.
6. Adjust the coarse focus knob, while looking into the eyepiece, until the
image you see is clear and in focus.
7.
Answers
1. The specimen on a slide is placed here on the stage and clipped into
place.
2. By rotating the nosepiece, the lowest power (smallest) objective lens is
placed over the specimen.
3. Adjust the coarse focus knob, while looking into the eyepiece, until the
image you see is clear and in focus.
4. Whilst viewing the image adjust the iris diaphragm for optimum light.
5. Make sure that the object you wish to view is directly over the hole in
the stage. Now rotate the nosepiece and bring the x10 objective into
place over the specimen. Look down the ocular tube and use the fine
focus knob to focus the image.
6. Repeat step 5 using the x40 objective lens.
Calculating Magnification
Total magnification =
magnifying power of the objective lens x magnifying power of the
eyepiece lens
An actual object is small. An image is much
larger
What is the magnification of this
microscope?

x2
5mm 10mm
What is the magnification of this
microscope?

x5
2mm 10mm
Photomicrograph
A photograph of the image seen using an optical microscope . Modern
digital microscopes display the image on a computer screen.
 ●
microscope ●
uses ●
advantages ●
disadvantages

●
Optical (light)
microscope

●
Electron microscopes (EM)

●
Transmission
EM

●
Scanning EM

●
Laser scanning
confocal
microscope
Optical Microscopes
• The development of optical (light) microscopes played a key role in
our understanding of cell structure. They are the main ones used in
schools. They are:
• Relatively cheap
• Easy to use
• Portable and able to used in the field as well as in laboratories
• Able to be used to study whole living specimens
•
Optical Microscopes
• Optical microscopes allow magnification up to x1500 or in some types
x2000, which enables us to see clearly some of the larger structures
inside cells.
• However, because their resolution is limited, they cannot magnify any
higher while still giving a clear image.
• Optical microscopes use visible light, a part of the electromagnetic
spectrum that has a wavelength of between 400 and 700nm.
• The wavelength of visible light ranges from 400 to 700nm so structures
closer together than 200nm will appear as one object.
• Ribosomes are very small, non-membrane-bound, cell organelles of about
20nm diameter, and so they cannot be examined using a light
microscope.
In the light microscope you can visualize the
outer boundary of the cell and the nucleus with
little detail in the cytoplasm
Electron Microscopes
• Electron microscopes use a beam of fast-travelling electrons with a
wavelength of about 0.004nm. This means that they have much
greater resolution than optical microscopes and can be used to give
clear and highly magnified images.
• The electrons are fired from a cathode and focused, by magnets
rather than glass lenses, on to a screen or photographic plate.
• Fast-travelling electrons have a wavelength about 125,000 times
smaller than that of the central part of the visible light spectrum.
This accounts for an electron microscope’s much better resolution
compared with an optical microscope.
Transmission electron
microscopes
• The specimen has to be chemically fixed by being dehydrated and
stained.
• The beam of electrons passes through the specimen, which is stained
with metal salts. Some electrons pass through and are focused on
the screen or photographic plate.
• The electrons form a 2D black and white (grey scale) image. When
photographed this is called an electron micrograph. Transmission
electron microscopes can produce a magnification of up to 2 million
times, and a new generation is being developed that can magnify up
to 500, 000.
Transmission Electron Microscope (TEM):
electrons pass through specimen
1. Electron gun
2. Condenser lens
3. Specimen plane
4. Objective lens
5. Projector lens system
6. Fluorescent screen
7. Intermediate image planes
8. Binocular viewer

Specimens must be thin, so electrons can pass through them

Scanning electron microscopes
• These were developed during the
1960s. Electrons do not pass
through the specimen, which is
whole, but cause secondary
electrons to ‘bounce off’ the
specimen’s surface and be
focused on to a screen. This
gives a 3D image with a
magnification from x15 up to
x200000. The image is black and
white, but computer software
programmes can add false
colour. However, the specimen
still has to be placed in a
vacuum and is often coated with
a fine film of metal.
Scanning Electron Microscope (SEM): electrons
bounce off surface to produce 3D appearance
Laser scanning microscopes
• Laser scanning microscopes are also called confocal microscopes.
• They use laser light to scan an object point by point and assemble, by computer, the
pixel information into one image, displayed on a computer screen.
• These images are high resolution and show high contrast.
• These microscopes have depth selectivity and can focus on structures at different
depths within a specimen. Such microscopy can therefore be used to clearly
observe whole living specimens, as well as cells.
• They are used in the medical profession, for example to observe fungal filaments
within the cornea of the eye of a patient with a fungal corneal infection, in order
to give a swift diagnosis and earlier and therefore more effective treatment.
• They are also used in many branches of biological research.
Both types of electron
microscope:
• Both types of electron microscope:
• Are large and very expensive
• Need a great deal of skill and training to use
•
• Specimens, even whole ones for use in SEMs have to be dead, as they
are viewed while in a vacuum.
• The metallic salt strains used for staining specimens may be
potentially hazardous to the user.
 ●
microscope ●
uses ●
advantages ●
disadvantages
●
Uses light rays to ●
Can observe living ●
Low magnification (up
observe object. things to 2000 times)
●
Does not use harsh ●
Low resolution
●
light chemicals
microscope ●
Easy to set up and use
●
Cheap and portable

●
Electron microscopes (EM)
●
Uses focused beams of ●
High magnification (up to ●
Can only see dead
electrons through 5000 000 times) material
●
Transmission sections of tissues. ●
High resolution ●
Harsh chemicals
EM ●
Can see details inside cells used in
preparation which
can cause artefacts
●
Uses focused beams of ●
High magnification (up to ●
Expensive
electrons reflected off 500 000 times)
the tissues. ●
High resolution
●
Scanning EM
●
Can see details of the
surfaces of structures
●
Uses a laser beam of ●
Can see living cells. ●
More expensive
light to illuminate ●
Can observe cell processes than light
chemical stains within by tracking molecules. microscope.
●
Laser scanning
the specimen. These Higher resolution than ●
More complex
microscope then fluoresce.
●

light microscopes. than light

microscope.
Exam Questions
 Units of measurement
• In biology the most common unit of measurement is a
micrometer (µm)
•
• Nano= billionth
• Micro = means millionth
• Milli = means 1000th
• Centi = means 100th
•
 Units of measurement
• In biology the most common unit of measurement is a
micrometer (µm)
•
• nm = nanometer = 1 x10-9m one billionth of a m
• µm = micrometer = 1x 10-6m millionth of a m
• mm= millimeters = 1 x10-3m thousandth of a m
• Cm = centimeter = 1 x 10-2m 100th of a m
 Units – useful for
microscopy
Unit Symbol Size in meters
Metre m =1m
Millimetre mm = 10-3 m
Micrometre µm = 10-6 m
Nanometre nm = 10-9 m
Picometre pm = 10-12 m
The international (SI) unit for length is the meter (m)
1 mm = 1000 μm
1 μm = 1000 nm
Micro in Greek means millionth
To convert µm into mm you have to divide by a 1000. for e.g.
250µm = 0.25mm (250 / 1000)
Practice
1.
2. convert 50mm to micrometers
3. Convert 6cm into micrometers
4. Convert 300 millimeters (mm) into meters
5. Convert 2mm into nanometers
•
•
•
÷1000 ÷1000
•
mm µm nm
x1000 x1000

X1000 000
Practice
1.
2. convert 50mm to micrometers
3. Answer: 50x 1000= 50,000µm
4. Convert 6cm into micrometers
5. Answer: 6cm = 6 x 10000= 60,000µm
6. Convert 300 millimeters (mm) into meters
7. Answer: 300÷1000 = 0.3m
8. Convert 2mm into nanometers
9. Answer: 2 x ÷1000
1 000 000 = 2 000
÷1000
000 nm
•
• mm µm nm
x1000 x1000
•
• X1000 000
Summary
●
Microscope ●
Resolution ●
Magnification
●
Light ●
200nm or ●
X1500 –
●
0.2µm x2000
●
Transmission ●
0.1-0.3nm ●
x300,000
electron
microscope
●

Scanning ●
A little over ●
X200,000
electron 0.5nm
microscope
●
• In a table summarise electron and light microscopes.
Relate to magnification, resolution, use etc.

●
Light ●
Electron
microscope microscope
Extension:
Suggest the most useful type of microscope to observe the following
with an explanation:
a) Living water fleas in pond during a biology field trip
b) Cells taken from a cervical smear to check for abnormal cells
c) Virus particles
d) Ribosomes in a liver cell
01/06/25

2.1.2 Slides and

photomicrographs
Do now (5 minutes):
1) Define magnification
2) Define resolution
3) Name an advantage of a light microscope
3)

STOP

Extension: Compare the differences between a transmission electron microscope and scanning electron
microscope
 Do now corrections
Do now corrections (in green pen)
Define magnification
How much larger an image appears in comparison to an original object
Define resolution
Ability to see detail finely and be able to distinguish as two separate objects
Name an advantage of a light microscope
Cheap/easy to use/portable
1)

Extension: Compare the differences between a transmission electron microscope and scanning electron microscope
TEM- Electron passes through specimen but for SEM they do not
 This lesson: Slides and photomicrographs

•
• To understand the preparation of slides
• To understand the uses of staining
• To calculate magnification and rearrange the
formula
•

This half term

This Unit – Cell structure 2.1

2.1.1- Microscopes
2.1.2- Slides and photomicrographs
2.1.3- Measuring objects seen with a light
microscope (PA)
2.1.4- The ultrastructure of eukaryotic cells
PAG 1.1- Microscopy
2.1.5- Other features of eukaryotic cells
2.1.6/2.1.7 (PWFT)
2.1 Revision
Converting units
●
Unit ●
How many
x1000 millimetres it ÷1000
is
x1000 ÷1000
●
Millimetre (mm) ●
1
●
Micrometre (µm) ●
0.001
2.
●
Nanometre
1. The diameter (nm)
of an arteriole 0.000001
is 1.5 mm, how● many µm is it?
3.
4.
5.
1.5  1000 = 1500 µm
6.
7.

8. A mitochondrion is 2 µm long, how many nm is it?

9.
10.

2  1000 = 2000 nm
11.
12.
13.
14.

15. A chloroplast is 10 500 nm, what size is it in µm?

10500/ 1000 = 10.5 µm
Magnification = Image size ( as observed)

Actual size of object

Q. Calculate the actual width of
the spider

White crab spider, magnification x3

42 mm actual width = image length ÷ magnification

width of spider in image = 42 mm

magnification x3

= 42 mm ÷ 3

= 14 mm

The spider is three times smaller than the

image.
 Q. Calculate the actual length of
the duck
African white-backed duck
(magnification x ¼)

actual width = image length ÷ magnification

width of duck in image = 64 mm

magnification x ¼

= 64 mm ÷ ¼
64 mm = same as 64 mm x 4

= 256 mm

The duck is four times bigger than the image.

Fritillary butterfly, magnification x ½
Calculate the wingspan of the butterfly

28mm / 0.5
=56mm
Rhopalid bug, magnification x3
Calculate the actual length of the bug

21mm / 3
= 7mm

Hereford cow, magnification x 1/20

Calculate the actual height of the cow

57mm / 0.05
= 1140mm
=1.14m
Grapes on a vine, magnification x ½
Calculate the actual length of the bunch of grapes

80mm / 0.5
= 160mm
=16cm

Slow worm, magnification x 1/3

Calculate the actual length of the worm
75mm / 0.33
= 225mm
= 22.5cm
More questions to try
1. Calculate the magnification if you have a magnified image that’s 5mm wide
and an object that is 0.05mm wide.
2.
3.
4.
5.

6. Calculate the image size if your specimen is 0.1mm wide and the
magnification is x20.
7.
8.
9.
10.

11. Calculate the object size if your magnified image is 5mm and the magnification
is x50
12.
 Think, turn & talk task

What are the steps to use this type of

microscope?

You can view:

- Living organisms like
amoeba
- Blood/cheek cells
- Thin sections like
bone/muscle/leaf
Observing unstained specimens

Many single celled organisms like paramecium are transparent, some microscopes use light
interference and not absorption
Why do we stain specimens
Why do we stain specimens
• Specimen are usually colourless, and cell parts need
to take up dyes so the structures can be seen more
clearly.
Preparing Specimens
• In a optical (light) microscope, we use thin slices of
animal or plant material
• These can be placed on a slide and flooded with
water
• A coverslip is used to cover this wet mount
 Stains
• Stains combine with certain chemicals inside the cells and
their colour shows where they are, for e.g. methylene
blue is used to stain nuclei
• Different cells will stain differently because they contain
different chemicals
 Rat intestine,
Lung tissue of an asthmatic
fluorescence
patient.
microscopy, using DAPI
for staining nuclei
(blue)
Differential staining
The stains involved in differential staining are:
• Acetic orcein
• Eosin
• sudan red
• Iodine in potassium iodide
• Acetic orcein – binds to DNA and
stains chromosomes red

• Sudan red – stains lipids

• Eosin – stains cytoplasm

• Iodine in potassium iodide

– stains cellulose in plant cell walls
yellow, and starch grains blue/black
Longitudinal section Vs transverse/cross section
Exam Questions
1. A student suggested the details of structure C (mitochondria) could
be seen clearly with a good light microscope. Explain why the
student is not correct (2 marks)
2. Staining is a process often used in microscopy. Describe the
advantages of staining specimens viewed under a microscope (2
marks)
•
Answers
1) Mitochondria is too small , resolution is not high enough / only 0.2
micrometers
2) Makes it visible/ easier to see/ more detail, increases/ provides
contrast, identify cell types/ organelles
•
PA task
Homework
• Watch a video on how to use a graticule for a microscope.
• Make notes and draw diagrams on this, this must be brought to next
lesson.
01/06/25

2.1.3 Measuring objects

with a light microscope
Do now (5 minutes):
1) What is placed on top of a slide to protect a specimen?
2) Name two types of stains
3) What is the function of mitochondria
4)

Challenge: Where are plasmids found and what is the shape of them
STOP
 Do
Do now corrections (in green pen)
now corrections
What is placed on top of a slide to protect a specimen?
Coverslip
Name two types of stains?
Sudan red/eosin
What is the function of mitochondria
Releases energy through aerobic respiration
1)

Challenge: Where are plasmids found and what is the shape of them
In bacteria and they are circular
 This lesson: Microscopes

•
• To understand the difference between
magnification and resolution
• To be able to describe different types of
microscopes
•

This half term

This Unit – Cell structure 2.1

2.1.1- Microscopes
2.1.2- Slides and photomicrographs
2.1.3- Measuring objects seen with a light
microscope (PA)
2.1.4- The ultrastructure of eukaryotic cells
PAG 1.1- Microscopy
2.1.5- Other features of eukaryotic cells
2.1.6/2.1.7 (PWFT)
2.1 Revision
(b) the preparation and examination of microscope slides for use in
light microscopy *Including the use of an eyepiece graticule and
stage micrometer

Key words: eyepiece graticule, stage graticule

•
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•
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Please remember your labcoats for Tuesday!

HW check: Notes on graticules
Cell biology quiz
1) What does the cell wall do?
2) What does the cell membrane do?
3) What does the nucleus do?
4) What does the mitochondria do?
5) What do ribosomes do?
6) What provides plants with the green pigment?
7) Name 3 organelles plant cells have that animal cells don’t
8) Where is the DNA in bacterial cells kept?
9) What structure allows some bacterial cells to move?
10) Compare the differences between eukaryotic and prokaryotic cells
The Eye Piece Graticule
• The graticule is a circular glass
or plastic disc which fits
inside the eye piece of a
microscope
• It has a scale engraved on it
surface, usually with 100
divisions, with the main
divisions numbered 1 to 10
• The scale is meaningless
without calibration to give it
equivalency to a μm
 Staged Micrometer/ graticule
• Placed on the microscope stage, and used to calibrate the value of eye piece
divisions at different magnifications.
• This is used to calibrate the eye piece graticule
• It is a glass slide with an accurate scale marked on it which is usually 1mm long
and has 100 divisions
• Each division is often equal 10μm (0.01mm)
 Calibration
• Remove specimens from
slide
• Place stage micrometer
slide on stage of
microscope
• Bring into sharp focus with
the lowest magnification
Ruler and calculator
 Calibration example
• Each division of the stage graticule is 0.01mm or 10µm;
• In this example 15 eyepiece units corresponds to 10 divisions of the stage micrometer
• 100 μm ( each division = 10 μm, so 10x10) / 15 = 6.67 μm
• So if a cell is 5 divisions long using the eye piece graticule, the size will be 5 x 6.67 =33.33µm
•
•
graticule

graticule
1. After inserting a stage graticule on the microscope stage, bring it into focus
using the lowest power objected ( x4)
2. Total magnification is now x 40 ( 10x4 –eyepiece lens magnification x objective
lens magnification)
3. Align eye piece graticule with stage graticule as shown below.
4. Work out the value of one eye piece division at this magnification
Calibrating at x 4/x40 using this example,
• The stage graticule has 100
divisions, (1mm long/1000µm)
40 eye piece divisions
• Here, 10 divisions on the stage
graticule corresponds to 40
eye piece divisions
• Each eye piece division is equal
to 10 µm, so 4 divisions = 40
µm
• The stage graticule is 100
divisions long, each division is
10 µm
• Therefore, 1 eye piece division =
1000 µm/40 =25 µm
•
Calibrating at x10/x100 using this example,
• Here, 10 divisions on the
10 divisions= 100 eye piece divisions eye piece graticule
40 eye piece divisions corresponds to 100
stage graticule
Eye piece graticule divisions
• Each eye piece division is
equal to 10 µm, so 10
divisions = 100 µm
• Therefore, 1 eye piece
division
= 1000 µm/100 =10 µm
•
100 divisions
Stage gaticule =1000 µm
• The cell is 7 lines wide when
observed on x10
• Size of cell (29-22)
• = 7 x 10µm ( eye piece
division) = 70µm
Each 1 division is = to 10 micrometres. A cheek
cell has a diameter of 18 units on the eyepiece
graticule scale. Calculate the actual diameter of
the cheek cell in micrometres.
Complete your worksheet
 You should now calibrate the objective lenses and
record your results in a table:
•

Objective value of 1 eye piece division

X4
X 10
X 40
 Biological drawings
• Draw outline
• Do not shade
or add detail
•
Plant and animal structure

•
List the differences in structure of plant and animal
cells.