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CIE AS Biology Your notes
1.1 The Microscope in Cell Studies

Contents
The Microscope in Cell Studies
Magnification Calculations
Eyepiece Graticules & Stage Micrometers
Resolution & Magnification
Calculating Actual Size
Page 1 of 25
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The Microscope in Cell Studies

Your notes
Microscope Slide Preparation
Preparing a microscope slide
Specimens can be viewed under a light microscope; this allows some details of cellular material to be
observed
Pre-prepared permanent slides can be viewed
Such slides are produced by cutting very thin layers of tissue which are stained and permanently
mounted on a glass slide for repeated use
Different methods will be used to view different types of specimen, e.g. temporary slide preparations
can be produced in the school laboratory as described below
Preparing a slide using a liquid specimen
1. Add a few drops containing the liquid sample to a clean slide using a pipette
2. Lower a coverslip over the specimen and gently press down to remove air bubbles
Coverslips protect the microscope lens from liquids and help to prevent drying out
Preparing a microscope slide using a solid specimen
1. Use scissors or a scalpel to cut a small sample of tissue, and peel away or cut a very thin layer of cells
from the tissue sample
The preparation method always needs to ensure that samples are thin enough to allow light to
pass through
2. Place the sample onto a slide
A drop of water may be added at this point
3. Apply iodine stain
4. Gently lower a coverslip over the specimen and press down to remove any air bubbles
Preparing a microscope slide using onion cells diagram
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Your notes
Tissue from an onion is as a solid specimen, and can be prepared here using iodine stain
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Preparing a slide using human cells

1. Brush teeth thoroughly with normal toothbrush and toothpaste Your notes
This removes bacteria from teeth so they don't obscure the view of the cheek cells
2. Take a sterile cotton swab and gently scrape the inside cheek surface of the mouth for 5-10 seconds
3. Smear the cotton swab on the centre of the microscope slide for 2-3 seconds
4. Add a drop of methylene blue solution
Methylene blue stains negatively charged molecules in the cell, including DNA and RNA
This causes the nucleus and mitochondria to appear darker than their surroundings
5. Place a coverslip on top
Lay the coverslip down at one edge and then gently lower the other edge until it is flat
This reduces bubble formation under the coverslip
6. Absorb any excess solution by allowing a paper towel to touch one side of the coverslip
Preparing a microscope slide using cheek cells diagram
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Your notes
Cheek cells can be stained using methylene blue

Staining specimens
The cytoplasm and other cell structures may be transparent or difficult to distinguish; stains allow them
to be viewed clearly under a light microscope
As with the type of preparation required, the type of stain used is dependent on the specimen being
viewed
Common microscope stains & uses table
Stain Uses
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Stains starch blue-black, and colours nuclei and plant cell

Iodine Your notes
walls pale yellow
Crystal violet Stains cell walls purple
Methylene blue Stains animal cell nuclei dark blue
Is not taken up by cells and stains the background red, so

Congo red
providing contrast with any cells present
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Drawing Cells
To record the observations seen under a microscope, a labelled biological drawing is often made Your notes
Biological drawings are line drawings which show specific features that have been observed when the
specimen was viewed
There are a number of rules/conventions that are followed when making a biological drawing
The drawing must have a title
The magnification under which the observations shown by the drawing are made should be
recorded if possible
A scale bar may be used
A sharp pencil should be used
Drawings should be on plain white paper
Lines should be clear, single lines without sketching
No shading should be used
The drawing should take up as much of the space on the page as possible
Well-defined structures should be drawn
Only visible structures should be drawn, and the drawing should look like the specimen
The drawing should be made with proper proportions
Structures should be clearly labelled with label lines that:
Do not cross
Do not have arrowheads
Connect directly to the part of the drawing being labelled
Are on one side of the drawing
Are drawn with a ruler
Drawings of cells are typically made when visualizing cells at a higher magnification power, whereas
plan drawings are typically made of tissues viewed under lower magnifications (individual cells are
never drawn in a plan diagram)
Plant cell biological drawing
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Bacterial cell biological drawings

Your notes
Page 8 of 25
Animal cell drawing

Your notes
Exam Tip
When producing a biological drawing, it is vital that you only ever draw what you see and not what you
think you should see!
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Your notes
Magnification is the number of times that a real-life specimen has been enlarged to give a larger
view/image
E.g. a magnification of x100 means that a specimen has been enlarged 100 times to give the image
shown
The magnification (M) of an object can be calculated if both the size of the image (I), and the actual size
of the specimen (A), is known
An equation triangle allows the magnification equation to be easily rearranged

When carrying out calculations relating to magnification, the following steps should be followed:
1. Rearrange the equation
A=I÷M
M=I÷A
I=AxM
2. Read and measure the relevant values from the question stem and/or any images provided
3. Convert any units
4. Substitute numbers into the rearranged equation
5. Consider whether this value makes sense in the context provided
Converting units during magnification calculations

Cellular structures are usually measured in either micrometers (μm) or nanometers (nm), while any
measurements carried out in an exam with a ruler are likely to be in millimeters (mm)
There are 1000 µm in a mm
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There are 1000 nm in a µm

When doing calculations all measurements must be expressed using the same units
It is best to use the smallest unit of measurement shown in the question Your notes
Note that magnification does not have units
To convert units, multiply or divide depending on whether the units are increasing or decreasing
Multiply when converting from a larger unit to a smaller unit
Divide when converting a smaller unit to a larger unit
Units can be multiplied or divided by a factor of 1000 when converting between mm and µm
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Worked example
Your notes
A starch grain inside a plant cell was viewed under a microscope at a magnification of x850. The image
of the starch grain captured using the microscope is shown below.
Calculate the actual diameter of the starch grain between points C-D in the image.
Give your answer in μm.
Step 1: Rearrange the equation
We have been asked to calculate actual size, A, so the new equation should be
Actual size = image size ÷ magnification
Step 2: Read and measure relevant values
We need to know the image size and the magnification
The image size has been given in the image as 20 mm
The magnification has been included in the question stem and is x850
Step 3: Convert any units
The actual diameter of a structure inside a cell would normally be measured in μm
The image size has been given in mm, so we need to convert this to μm
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1 mm = 1000 μm, so we multiply mm by 1000 to give the value in μm

20 x 1000 = 20 000 μm Your notes
Step 4: Substitute number into the equation
Actual size = 20 000 ÷ 850
= 23.53 μm
Step 5: Consider whether the value makes sense
Plant cells measure between 10-100 μm, and the starch grain takes up a large proportion of the cell in
the image. A diameter of around 23 μm could therefore be right for this starch grain
If we had forgotten to convert the units and come up with a value of 0.024 μm, then this step would
show us that this is probably too small and that we must have therefore made a mistake somewhere
Exam Tip
Magnification calculations will usually involve measuring images on a page, so you must have a ruler
with you in an exam!
Be aware that the unit conversions given above only work if the initial measurement is taken in mm,
not in cm. Any measurements taken in cm will need to be converted into mm before multiplying by
1000 to give μm.
Some exam questions may expect you to use a scale bar to calculate the magnification of an image.
Remember that you can use a scale bar to find both actual size (written on the scale bar) and image size
(the measured length of the bar), so these values can be used to calculate magnification using the
equation M = I ÷ A
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Your notes
An eyepiece graticule and stage micrometer are used to measure the size of an object when viewed
under a microscope
The eyepiece graticule is an engraved ruler that is visible when looking through the eyepiece of a
microscope
Eyepiece graticules are often divided into 100 smaller divisions known as graticule divisions, or
eyepiece units
The values of the divisions in an eyepiece graticule vary depending on the magnification used, so the
graticule needs to be calibrated every time an object is viewed
The calibration is done using a stage micrometer; this is a slide that contains a tiny ruler with an accurate
known scale
Stage micrometer rulers can vary, but often have larger divisions of 0.1 mm (100 μm) and smaller
divisions of 0.01 mm (10 μm)
Calibrating the eyepiece graticule
In the diagram, two stage micrometer divisions of 0.1 mm, or 100 μm, are visible
Each 100 µm division is equal to 40 eyepiece graticule divisions
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40 graticule divisions = 100 µm

1 graticule division = number of µm ÷ number of graticule divisions
Your notes
1 graticule division = 100 ÷ 40 = 2.5 µm; this is the magnification factor
Calculating the size of a specimen
The calibrated eyepiece graticule can be used to measure the length of an object
The number of graticule divisions covered by an object need to be multiplied by the magnification
factor:
graticule divisions covered by object x magnification factor = length of object (µm)
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Worked example
Your notes
A student viewed some onion cells under a microscope.
The image below shows the cells with an eyepiece graticule (left) and the eyepiece graticule alongside
a stage micrometer (right).
Note that each large division on the stage micrometer here is 100 μm, and each small division is 10 μm.
Use the stage micrometer to calibrate the eyepiece graticule and calculate the actual length of the cell
labelled C-D in the image.
Step 1: Calculate the size of each eyepiece division
There are 40 graticule divisions per large micrometer division, or per 100 μm
1 graticule division = no. of μm ÷ no. of graticule divisions
= 10 ÷ 40
= 2.5 μm
This value can now be used as a magnification factor
Step 2: Calculate the length of the cell
Specimen size = no. of graticule divisions x magnification factor
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The cell closest to the ruler covers 27 graticule divisions

= 27 x 2.5 Your notes
= 67.5 μm
Step 3: Consider whether this answer makes sense in context
Plant cells usually measure between 10-100 μm, so a result of 67.5 μm sounds sensible in this context.
Exam Tip
The calculations involving stage micrometers and eyepiece graticules are often seen in exam
questions, so make sure that you are comfortable with how to calibrate the graticule and calculate the
length of an object on the slide.
Note that both of the examples given above are carried out at the same magnification, so the
magnification factor calculated during calibration is 2.5 in both cases. This may not always be the same
in an exam.
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Your notes
Magnification
Magnification is the number of times that a real-life specimen has been enlarged to give a larger
view/image
E.g. a magnification of x100 means that a specimen has been enlarged 100 times to give the image
shown
A light microscope has two types of lens which allow it to achieve different levels of magnification:
An eyepiece lens, which often has a magnification of x10
A series of objective lenses, each with a different magnification, e.g. x4, x10, x40 and x100
To calculate the total magnification of a specimen being viewed, the magnification of the eyepiece
lens and the objective lens are multiplied together:
total magnification = eyepiece lens magnification x objective lens magnification
Resolution
The resolution of a microscope is its ability to distinguish two separate points on an image as
separate objects; this determines the ability of a microscope to show detail
If resolution is too low then two separate objects will be observed as one point, and an image will
appear blurry, or an object will not be visible at all
The resolution of a microscope limits the magnification that it can usefully achieve; there is no
point in increasing the magnification to a higher level if the resolution is poor
The resolution of a light microscope is limited by the wavelength of light
Visible light falls within a set range of light wavelengths; 400-700 nm
The resolution of a light microscope cannot be smaller than half the wavelength of visible light
The shortest wavelength of visible light is 400 nm, so the maximum resolution of a light
microscope is 200 nm
E.g. the structure of a phospholipid bilayer cannot be observed under a light microscope due to
low resolution:
The width of the phospholipid bilayer is about 10 nm
The maximum resolution of a light microscope is 200 nm, so any points that are separated by a
distance of less than 200 nm, such as the 10 nm phospholipid bilayer, cannot be resolved by a
light microscope and therefore will not be distinguishable as separate objects
Electron microscopes have a much higher resolution, and therefore magnification, than light
microscopes as electrons have a much smaller wavelength than visible light
Electron microscopes can achieve a resolution of 0.5 nm
Resolution of light and electron microscopes diagram
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Your notes
The resolving power of electron microscopes is much greater than that of light microscopes due to the
smaller wavelength of electrons in comparison to visible light
Comparison of light and electron microscopes
Light microscopes are used for specimens larger than 200 nm
Light microscopes shine light through the specimen
The specimens can be living, and therefore can be moving, or dead
Light microscopes are useful for looking at whole cells, small plant and animal organisms, and
tissues within organs such as in leaves or skin
Electron microscopes, both scanning and transmission, are used for specimens larger than 0.5 nm
Electron microscopes fire a beam of electrons at the specimen
Transmission electron microscopes (TEM) fire electrons through a specimen
Scanning electron microscopes (SEM) bounce electrons off the surface of a specimen
The electrons are picked up by an electromagnetic lens which then shows the image
Electron microscopy requires the specimen to be dead, meaning that they can only be used to
capture a snapshot in time, and not active life processes as they occur
Electron microscopes are useful for looking at organelles, viruses and DNA as well as looking at
whole cells in more detail
Comparing light & electron microscopes table
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Electron microscope Light microscope

Your notes
Large machines that are permanently installed in
Small and portable
laboratories
Need to create a vacuum for electrons to travel
No vacuum required
through
Specimen preparation is complex Specimen preparation can be simple
Maximum magnification of x500 000 Maximum magnification of x2000
Maximum resolution of 0.5 nm Maximum resolution of 200 nm
Specimens are always dead Specimens can be living or dead
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Your notes
When using microscopes to study biological specimens, it is possible to calculate the actual size of
organisms, cells, and parts of cells
The actual size of specimens can be calculated using the magnification and the measured size of an
image as follows:
Magnification is sometimes provided in an exam question stem
Magnification can be calculated from the scale bar of an image
Sometimes magnification is calculated from information about the magnification of the eyepiece
lens and the objective lens
Remember that the equation above is part of the equation triangle from calculating magnification
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Worked example
Your notes
Using a scale bar to calculate actual size
A lab technician observed bacterial cells with an electron microscope, and produced the image
below.
The scale bar measures 2 cm in length, and the length of the technician's image of one bacterial cell
measures 7.6 cm.
Use the information provided to calculate the actual length of a bacterial cell in the image.
Step 1: Use the scale bar to calculate the magnification of the image
The equation triangle for magnification tells us that:
Magnification = image size ÷ actual size
The scale bar measures 2 cm = 20 mm = 20 000 μm
The scale bar represents an actual size of 1 μm
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Magnification = 20 000 ÷ 1
= 20 000 Your notes
Step 2: Substitute values into the equation for actual size
The question stem tells us that one cell = 7.6 cm = 76 mm = 76 000 μm
Magnification is ×20 000
Actual size = 76 000 ÷ 20 000
= 3.8
Therefore, the actual length of a bacterial cell in this image is 3.8 μm
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Worked example
Your notes
Using lens magnification to calculate actual size
A scientist looked at a sample of red blood cells under a light microscope.
The eyepiece lens of the microscope had a magnification of ×10 and the objective lens had a
magnification of ×40.
The scientist produced a photomicrograph of the blood cells, shown below, in which the red blood
cells have an average diameter of 3 mm when measured using a ruler.
Calculate the average diameter of the red blood cells in the sample. Give your answer in micrometres.
Step 1: Calculate the total magnification of the specimen
total magnification = eyepiece lens magnification × objective lens magnification
10 × 40 = 400
Magnification = ×400
Step 2: Convert the image size into μm
1 mm = 1000 μm
3 × 1000 = 3000
Image size = 3000 μm
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Step 3: Substitute values into equation for actual size

Actual size = image size ÷ magnification Your notes
Actual size = 3000 ÷ 400
= 7.5
Therefore, the average size of a red blood cell in this sample is 7.5 μm
Exam Tip
Note that you would be expected to use your own ruler to provide image size measurements in actual
size calculations.
Don't forget to convert units in calculations like this!
Page 25 of 25

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CIE AS Biology Your notes

1.1 The Microscope in Cell Studies

The Microscope in Cell Studies

Preparing a slide using human cells

Cheek cells can be stained using methylene blue

Stains starch blue-black, and colours nuclei and plant cell

Crystal violet Stains cell walls purple

Methylene blue Stains animal cell nuclei dark blue

Is not taken up by cells and stains the background red, so

Bacterial cell biological drawings

Animal cell drawing

An equation triangle allows the magnification equation to be easily rearranged

Converting units during magnification calculations

There are 1000 nm in a µm

1 mm = 1000 μm, so we multiply mm by 1000 to give the value in μm

Eyepiece Graticules & Stage Micrometers

Calibrating the eyepiece graticule

40 graticule divisions = 100 µm

The cell closest to the ruler covers 27 graticule divisions

Resolution & Magnification

Electron microscope Light microscope

Specimen preparation is complex Specimen preparation can be simple

Maximum magnification of x500 000 Maximum magnification of x2000

Maximum resolution of 0.5 nm Maximum resolution of 200 nm

Specimens are always dead Specimens can be living or dead

Calculating Actual Size

Step 3: Substitute values into equation for actual size

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