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NotesBiologyTopic 1.4Microscopy skills
Back to Biology Topics
1.4.27 min read

Microscopy skills

IB Biology · Unit 1

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Contents

  1. 1Seeing cells: why we need microscopes
  2. 2Making a temporary mount
  3. 3Focusing the microscope
  4. 4Measuring with an eyepiece graticule
  5. 5Magnification, actual size and scale bars
  6. 6Exam-style question (step by step)
The big idea: Most cells are far too small to see with the naked eye. To study them we use a microscope — a tool that makes small objects look much bigger.

Two things matter when choosing a microscope

  • How much it magnifies (makes things look bigger).
  • Its resolution (how much fine detail it can show).
Microscope
A tool that produces an enlarged image of a small object so we can study it.
Magnification
How many times bigger the image looks than the real object.
Resolution
The ability to show two close points as separate, rather than as one blur — in short, how much fine detail can be seen.
Magnification is not the same as resolution: You can magnify a blurry image as much as you like — it just gets bigger and blurrier.

Resolution is what actually lets you make out new detail. A good microscope needs both.

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The big idea: To see cells with a light microscope, the specimen must be thin, flat and wet, and often stained.

The points to remember

  • Use a thin layer of tissue, one cell thick, so light can pass through it.
  • Lay it flat on a glass slide and add a drop of water or a stain.
  • Lower a cover slip at an angle, from one edge, so no air bubbles are trapped.
  • A stain colours some structures more than others, so they can be seen (iodine stains starch and nuclei).
Remember it as: Thin, wet, covered: no bubbles.
Side views: a thin layer peeled from an onion scale leaf; the layer on a slide with a drop of stain; a cover slip lowered at an angle with a needle; the finished flat mount
A temporary mount in four moves.
Real example: onion skin: The thin inner skin of an onion scale leaf is one cell thick. Peeled off and stained with iodine solution, its cell walls and nuclei show clearly.

When asked why a stain is used, name what it lets you see: for a root tip, the chromosomes, not just 'the cells'.

Making the mount is not viewing it: When asked how to prepare a mount, give the making steps only; putting it on the stage and focusing are not part of the preparation.

The cover slip is a thin piece of glass, not a second slide.

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The big idea: Find the specimen at low power, then focus in two steps: coarse first, fine second.

The points to remember

  • Start with the low-power objective lens: the field of view is larger, so the specimen is easier to find.
  • Focus with the coarse adjustment knob first, then sharpen with the fine adjustment knob.
  • Move to a higher power only once the part you want is in the centre of the view; refocus with the fine knob.
  • Total magnification = eyepiece magnification × objective magnification (×10 eyepiece × ×40 objective = ×400).
Remember it as: Low power to find it, fine knob to sharpen it.
1

Low power

Large field of view: the specimen is easy to find.

2

Coarse focus

Big movements bring the image roughly into focus.

3

Fine focus

Small movements make the detail sharp.

4

Higher power

Centre the part first, then refocus with the fine knob.

Real example: a school microscope: A typical school microscope has a ×10 eyepiece and objectives of ×4, ×10 and ×40, so its total magnifications are ×40, ×100 and ×400.
The big idea: A graticule turns 'it looks small' into a number: measuring with an instrument is a quantitative observation.

The points to remember

  • An eyepiece graticule is a small scale inside the eyepiece, used to measure cells.
  • Its divisions have no unit until it is calibrated with a stage micrometer, a slide with a scale of known length.
  • One graticule division = length on the stage micrometer ÷ number of graticule divisions it covers.
  • Actual size = number of divisions × the length of one division. Calibrate again at each objective lens.
An eyepiece scale from 0 to 100 above a stage micrometer whose 100 µm divisions each cover 25 graticule divisions; a cell 30 divisions long, 120 µm
Calibrate first, then count divisions.
The two lines of working: One division = 100 µm ÷ 25 = 4 µm.

Actual length = 30 × 4 µm = 120 µm.

A number can be compared, averaged and checked by someone else, which a description such as 'small' cannot. That is why measurements make observations more objective.

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The big idea: Magnification links three things: image size, actual size and magnification. Know two, find the third.

The points to remember

  • Magnification = image size ÷ actual size. Rearranged: actual size = image size ÷ magnification.
  • Put both lengths in the same unit first: 1 mm = 1000 µm.
  • A scale bar shows how long a length on the image is in real life. Magnification = scale bar's printed length ÷ the length written on it.
  • To draw a scale bar: printed length = actual length × magnification (10 µm at ×1000 = 10 mm).
  • Give the answer with a unit (µm) or a × sign. A photo through the eyepiece can be measured the same way.
Remember it as: I AM: Image = Actual × Magnification.
Onion epidermis cells stained with iodine; a red line across cell X and a 50 µm scale bar; printed lengths: scale bar 20 mm, cell X 48 mmOnion epidermis cells stained with iodine; a red line across cell X and a 50 µm scale bar; printed lengths: scale bar 20 mm, cell X 48 mm
Printed lengths are given under the image.
1

Change to the same unit

Scale bar 20 mm = 20 000 µm.

2

Magnification

20 000 µm ÷ 50 µm = ×400.

3

Actual size of cell X

48 mm = 48 000 µm; 48 000 ÷ 400 = 120 µm.

Units and the × sign: Change millimetres to micrometres before dividing. An actual size needs its unit (µm); a magnification needs a × and no unit.
How it is tested: Paper 1B opens with a micrograph: calculate a magnification, an actual size or a scale bar [1] each, name the eyepiece graticule, or describe how to make a temporary mount [3].
Three human cheek cells; a red line across cell X; a 20 µm scale bar; printed lengths: scale bar 22 mm, cell X 66 mmThree human cheek cells; a red line across cell X; a 20 µm scale bar; printed lengths: scale bar 22 mm, cell X 66 mm
The figure for this question.
IB-style questionCalculate[2 marks]

The light micrograph shows human cheek cells. On the printed page the scale bar is 22 mm long and the red line across cell X is 66 mm long.

(a) Calculate the magnification of the image.

(b) Calculate the actual width of cell X, giving the unit.

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For a temporary mount [3]: Three making steps, one per line: thin layer on a slide; a drop of water or stain; a cover slip lowered at an angle to avoid air bubbles.

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Yeast cells were viewed with a light microscope at a magnification of ×1000.

how the magnification of the microscope was calculated.
[1 mark]

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