2.1.

1a-f Microscopy (p8-25)

2.1.1a: the use of microscopy to observe and investigate different types of cells
and cell structure in a range of eukaryotic organisms

1. Describe, in principle, what a microscope does and name 4 different types of

microscopes. (F)
A microscope magnifies an image, making things easier to see that one would struggle
to see with the naked eye. Types of microscopes = SEM, TEM, Electron (light)
microscope, laser scanning confocal.

2. Label a diagram of a light microscope with the names of the component parts and
annotate this diagram with the functions of these parts.

3. State what “SEM” and “TEM” are abbreviations for. (F)

SEM – Scanning electron microscope
TEM – Transmission electron microscope

4. Outline how an SEM works.

An SEM produces a largely magnified image by using an electron beam that is directed
onto a sample. The electrons don't pass through specimen but bounce off. Reflected
electrons are then collected to form an image.

5. Outline how a TEM works.

A TEM uses a similar principle to the SEM, but SEM creates an image by detecting
reflected or knocked-off electrons, while TEM uses transmitted electrons (electrons that
are passing through the sample) to create an image. The electron beams pass through
a very thin sample. The electrons that pass through denser parts less easily gives
contrast.

6. Outline how a laser scanning confocal microscope works.

A single spot of light is moved across the sample (point illumination) causing
flourescence from 'dyed' parts. Light from the specimen is filtered through a pinhole so
only light from the focal plane giving the sharpest focus is detected.
 7. Draw a table comparing the use and properties of light, SEM, TEM, and laser scanning
confocal microscopes. (F)

Name Radiation Magnificatio Resolution Vacuu State of Stains

Used n m specimens
Light Light X1500-2,000 200nm No Alive or Contrast and
dead (thin) distinguish
features
SEM Electrons X100,000- 3-10nm Yes Dead Thin layer of
500,000 metal
TEM Electrons X500,000- 0.2-0.5nm Yes Dead Heavy metal ions
2,000,000 (thin) (contrast)
LSC Laser No limit 200nm Fluorescent dyes
light

8. State the features of the images produced from light, SEM, TEM and laser scanning
confocal microscopes.

Name Background Colours Details What is observed?

Light Light Stains/specimen 2D (limited depth) Large organelles
SEM 3D (large depth) Organelles within
cells on insects (e.g.)
TEM Grey (can have a 2D (small structures Smaller organelles.
false colour) can be grainy)
LSCM Black Only light collected Appears 2D but can Whole cells and
from flourescence. be processed to be stained structures
3D within cells.

9. Suggest how the development of the microscope influenced scientific thinking and what
the wider consequences of this might have been. (S+C)
Allowed cells to be observed - disproved spontaneous generation. Now it allows
theories to be presented then later disproved and replaced as technology improves.

2.1.1b: the preparation and examination of microscope slides for use in light
microscopy

1. Explain how to use a light microscope to view a specimen at low and high powers.
Place slide on stage and secure with stage clips
View through the scanning objective lens
Use the coarse adjustment to focus
Move to the next lens and re-focus
Move to the high-power lens and use fine focus to gain a clear image

2. Describe how to produce a temporary wet mount of living tissue.

Specimens are suspended in a liquid such as water or an immersion oil. A cover slip is
placed on from an angle.

3. Describe and explain the characteristics of a good slide preparation.

No artefacts, thin, transparent, clean slides & coverslips.
Good contrast, cells spread out with limited overlap.
Appropriate volume of liquid.

4. Explain why slide preparations need to be thin. (F)

To allow light through, and so that individual cells can be seen.
 5. Explain how to use a stage micrometre to work out the distance represented by the
small divisions in an eyepiece graticule under 3 different objective lenses. (F)
1) Line up eye piece graticule scale with stage units. (1 stage units = 0.1mm)
2) Divide the number of stage units (converted into mm) by the number of eyepiece
units they correspond to find the length of 1 epu.
3) Repeat for other lenses.
4) Size of each division of the eye piece graticule =distance on stage
micrometre/number of divisions of eye piece graticule

6. Explain how to use a stage micrometre and eye-piece graticule to add a scale bar to a
drawing.
Calculate the magnification of the drawing. The scale bar should be the length of
magnification x length it represents

7. Explain how to use a stage micrometre and eye-piece graticule to calculate the size of
a specimen.
Size of each division of eye-piece graticule x number of divisions = actual size of object

8. Describe how to choose an appropriate number of significant figures, or decimal places

to present data.
Use the highest number of decimal places in the question for your answer.

9. Explain how an adjustment to the “plane of focus” can alter what is viewed within a
cell. (F)
The microscope only focuses on a thin optical slice at any one time. As a cell is 3D,
different objects lie in different focal planes so can be brought in and out of focus.

10. Explain how a tissue slice might be misleading due to the very thin nature of the slice.
It is only a thin section of the whole tissue. Cells may appear to have no nucleus if it
was outside the section

2.1.1c: the use of staining in light microscopy.

1. Explain why staining is useful for light microscopy. (F)

Many specimens are transparent. Staining creates a contrast in colour or intensity
between objects and their surroundings, allowing them to be seen and identified.
Several different colours can be used on one specimen as they bind to different
molecules, so allow the molecules to be identified as present or absent.

2. Describe the properties a stain needs to have to be useful for light microscopy. (F)
Differential staining - must stain some objects and not others so that contrast is visible
Intense colour - only a very small amount of biological material but it must be visible

3. Describe how to prepare a stained specimen for viewing under a light microscope.
Fixing - preserve specimens in as near natural state as possible
Sectioning - sliced very thinly.
Staining - multiple stains for different structures
Mounting - secured to slide, coverslip on top, sealed to create permanent slide

4. Name two common stains and the molecules they bind to. (S+C)
Toluidine Blue - Phloem=red, Xylem=blue, DNA= blue
Iodine in potassium iodide - Starch=blue-black
2.1.1d: the representation of cell structure as seen under the light microscope using
drawings and annotated diagrams of whole cells or cells in sections of tissue.

1. State the rules for biological drawings. (F)

Sharp pencil
Clear continuous lines
No shading
Accuracy
More than half the available space
Correct mistakes with a good rubber
Include a title
Include a scale

2. Produce a labelled and annotated low power tissue plan using a light microscope.

3. Produce a labelled and annotated high power drawing of a named number of cells using
a light microscope.

2.1.1e: the use and manipulation of the magnification formula.

1. State the magnification formula
Magnification = Image size/ Actual size.

2. Explain how to calculate the magnification of an image using the magnification formula.
Measure image size, make sure image and actual are in the same units Divide image
by actual.

3. Explain how to calculate the actual size of an object using the magnification formula.
Image Size ÷ Magnification. Answer is in units of image size, may need converting.

4. State the symbols used for millimetres, micrometres, and nanometres. (F)
millimetres - mm
micrometres - μm
nanometres – nm

5. Explain how to convert measurements from one unit into another.

mm ×1000 → μm ×1000 → nm

6. Explain how to represent numbers in standard form.

Number between 1 and 10. Multiply by 10 to the power of x.
2.1.1f: the difference between magnification and resolution.

1. Define the terms “resolution” and “magnification”. (F)

Resolution = The ability to see two objects that are close together as separate objects
allowing detail to be seen.
Magnification = The number of times larger an image is compared to the object, of the
degree of enlargement of an image.

2. State the difference between magnification and resolution.

Magnification is enlargement but not necessarily increased detail. Magnification is just
a number -no units. Resolution is stated as distance - the smallest distance between
two objects where they can still be identified as separate

3. State the resolution and useful maximum magnification of light microscopes, SEMs and
TEMs.
Light microscope- 200nm, x1500-2000
SEM- 3-10nm, x100,000 -500,000
TEM- 0.2-0.5nm, x500,000 - 2,000,000

2.1.1g-k Cell structure (p26-37)

2.1.1g: the ultrastructure of eukaryotic cells and the functions of the different
cellular components

1. Define the terms “eukaryotic cell” and “ultrastructure”. (F)

Eukaryotes = any cell or organism that possesses a clearly defined nucleus
Ultrastructure = biological structure and especially fine structure (as of a cell) not
visible through an ordinary microscope.

2. Draw diagrams of a typical plant and animal cell, labelling the structures of the cell and
annotating with their function.

3. Draw and label a diagram of a mitochondrion, and a diagram of a chloroplast.

 4. State 5 similarities and 3 differences between a typical plant and animal cell.

Both have = nucleus, cell plasma membrane, cytoplasm, mitochondria, ribosomes.

Plant cells only = vacuole, chloroplasts, cell wall.
5. Draw a diagram showing the relative sizes of different cellular components. (S+C)
Nucleus - 3-20 micrometres
Chloroplast - 4-10 micrometres
Mitochondria - 2-5 micrometres
Lysosome - 0.5 micrometres
Centriole - 0.2 micrometres
Ribosome (80S) - 22nm
Ribosome (70S) - 18nm
Plasma Membrane - 5-7 nm

2.1.1h: photomicrographs of cellular components in a range of eukaryotic cells.

1. Identify organelles from images produced by light microscopy, TEM and SEM. (F)
TEM - transmission electron microscope (2D photos)
SEM - scanning electron microscope (3D photos)

2.1.1i: the interrelationship between the organelles involved in the production and
secretion of proteins.
1. Draw a flow chart that shows how different organelles and molecules are involved in
the process of protein production and trafficking in a cell. (F)
mRNA transcription —> ribosome translation —> vesicles transport —> Golgi
apparatus (modification) —> vesicles transport (exocytosis or used in cell)

2.1.1j: the importance of the cytoskeleton

1. Outline the structure of the 3 components of the cytoskeleton. (F)
2. Microfilaments allow cells to move by changing their shape and form a contractile ring
around the centre of the cell for cytokinesis
Microtubules form a scaffold to maintain overall cell shape and form tracks along which
organelles and motor proteins can move, as wells as forming cilia and flagella and
pulling chromosomes apart as centrioles
Intermediate filaments anchor organelles in place and maintain cell shape

3. Describe the functions of the cytoskeleton in a cell. (F)

Strengthening and support = providing mechanical strength to maintain strength and
keep the organelles in position.
Intracellular movement = forming ‘tracks’ along which organelles can move. (eg the
movement of vesicles.
Cellular movement = via cilia and flagellum. These both contain microtubules that are
responsible for moving them.

4. Describe the importance of the cytoskeleton in movements of chromosomes, cilia,

flagella, and vesicles.
In cell division the centrioles (microtubules) form the spindle, on which chromosomes
are separated.
Most flagella and mobile cilia are composed of 9 pairs of microtubules arranged in a
circle, along with an additional two microtubules in the centre of the ring. The body
made of microtubules keeps them in position
Microtubules allow vesicles to move within the cell so the contents of them can be
transported.

5. Describe the importance of the cytoskeleton in neutrophils.

 Cytoskeleton allows for flexibility in cell shape for phagocytosis/squeezing though small
gaps to get to site of infection as well as assisting in rapid cell division by aiding cytokinesis.
2.1.1k: the similarities and differences in the structure and ultrastructure of
prokaryotic and eukaryotic cells.

1. Define the term “prokaryotic cell”. (F)

A single-celled organism that has no true nucleus or membrane bound organelles. They
reproduce through binary fission and are unicellular.

2. List examples of both prokaryotic and eukaryotic cells.

Prokaryotic = bacteria, cyanobacteria.
Eukaryotic = palisade cell, red blood cell, lymphocytes.

3. Draw a diagram of a prokaryotic cell, label the structures, and function.

4. Draw a table outlining the structure and function of the cell wall, ribosomes, bacterial
flagellum, plasma membrane, plasmid, bacterial chromosome, cytoskeleton, pili and
slime capsule in prokaryotic cells. (F)

Cell Wall – rigid outer peptidoglycan covering maintaining cell structure.

Plasma membrane – lipid bilayer allowing selective substances such as sugar to pass
through.
Slime capsule – protective slimy layer to retain moisture and resist phagocytosis.
Plasmid – circular DNA
Pili – hair-like structure that attaches to other cells.
70S ribosomes – smaller than eukaryotes, site of protein synthesis.
Cytoskeleton – maintains shape and allows organelles to travel.
Bacterial chromosome – supercoiled DNA needed for protein synthesis.

5. Draw table comparing eukaryotic and prokaryotic cells. (F)

Eukaryotic = mitochondria, nucleus, sometimes cell wall, cytoplasm, larger
Prokaryotic = no mitochondria, no true nucleus, cell wall, cytoplasm, smaller
(There are more).

6. Describe the endosymbiotic theory. (S+C)

Some organelles within cells (mitochondria and chloroplasts) were once free-living
bacterial cells as evidenced by their own DNA and means for protein synthesis. At
some point they were ingested by a large host cell and became dependent on one
another, resulting in a permanent relationship. Through evolution, they became more
specialised and today they cannot live outside of the cell.