CALIBRATING EYEPIECE

GRATICULE USING
MICROMETER / CALCULATING
LINEAR MAGNIFICATION

Learning objectives: measure cell size using a microscope;

calculate the linear magnification using a micro photo;
KEY WORDS
• Magnification
• Actual size
• Image size
• Eyepiece graticule - a small piece of glass with a measurement scale on its
surface that fits inside a microscope eyepiece
• Stage micrometer - a microscope slide with
a scale etched on the surface
• Magnification factor
• Eyepiece lens
• Objective lens
CONVERTING UNITS OF
MEASUREMENT
 USING STAGE MICROMETER &
EYEPIECE GRATICULE
0.1 mm = 100 µm

40 graticule divisions = 100 µm

mm

1 graticule division = number of micrometres ÷

number of graticule division

1 graticule division = 100 ÷ 40 = 2.5 µm

This is the magnification factor

graticule divisions x magnification factor = measurement (µm)

CALCULATING ACTUAL SIZE

Known values: Solution:

• Eyepiece lens magnification:
x10
• Objective lens magnification:
x40
• Image size: 3 mm
eyepiece lens magnification x objective lens magnification
= total magnification
x10 x x40 = x400
3 mm = 3000 μm
RESOLUTION is the ability to distinguish between two separate points

• It’s limited by the wavelength of light Characteristics

• The longer the wavelength of light,

the more it is diffracted and the more
that this diffraction will overlap as the
points get closer together
• Electron microscopes have a much
higher resolution & magnification
than a light microscope as electrons
have a much smaller wavelength than
visible light
MCQ 1
A stage micrometer with small divisions of 0.1 mm was used to calibrate a graticule.
This is shown in the diagram below.
The stage micrometer scale is replaced by a slide of a plant cell.

What is the width of a chloroplast?

A) 0.5 mm В) 10 μm С) 50 μm D) 100 μm
The electronmicrograph shows a cell. MCQ 2
What is the actual diameter of the nucleus?

A) 0.6 μm
3.42 cm
В) 6 μm

C) 35 μm

D) 350 μm
MCQ 3
The diagram shows a graduated slide, with divisions of 0.1 mm viewed using an eyepiece
graticule. A
MCQ 4 D
MCQ 5
C
SQ 1
A student used a light microscope to view a specimen. They wanted to work out the actual
size of the specimen, and decided to use a stage micrometer, together with the ruler visible
inside the eyepiece of the microscope.

a) Outline why a stage micrometer is needed as well as the eyepiece graticule. [2]

• The size of the units of the eyepiece graticule change depending on the magnification being used; [1 mark]
• The stage micrometer can be used to calibrate the graticule; [1 mark]

b) Describe the steps required to calibrate the eyepiece graticule of a light microscope using
a stage micrometer. [3]

• Insert the eyepiece graticule and line up the scale of the graticule with the scale on the stage micrometer; [1 mark]
• Count the number of subdivisions of the stage micrometer that fit into each division of the eyepiece graticule; [1 mark]
• Multiply that number by the distance between each subdivision of the stage micrometer to give the distance between each division of the eyepiece graticule; [1 mark]
c) Explain why the steps outlined in part b) would need to be repeated when the student
changes to a more powerful objective lens. [2]

• (A change/increase in magnification) decreases the width of the field of view; [1 mark]

• As a result, the stage micrometer / specimen appears closer but the eyepiece graticule scale does not OR the specimen is zoomed in, taking up more of the field of view; [1 mark]

d) Fig. 3.1 shows the appearance of the scale of a stage micrometer. The length of the whole
scale is 1.00 mm. Point A shows the point at which the edge of a human cheek cell appears
in the microscope display. A typical cheek cell has diameter of approximately 50 μm.
Indicate on Fig. 3.1, with an arrow labelled B, the position of the opposite edge of the cheek
cell. [1]
SQ 2
Fig. 1.1 is an electron micrograph of part of a eukaryotic cell.

a) Describe the evidence from Fig. 1.1 that shows that

the image is a transmission electron micrograph.

Any 2 from
• (section at) high resolution; Accept suggestion of
a correct value of resolution for a TEM
• any named structure visible in Fig. 1.1 that can,
only be seen in a TEM / not be seen in a
photomicrograph; e.g. golgi bodies, ribosomes
• high magnification / higher magnification
(magnification > 1000 / higher than with light
microscope); in context of higher than light
microscope
b) State how it is possible to deduce that Fig. 1.1 is a transmission electron micrograph
and not a scanning electron micrograph. [1]
assume in context of transmission electron micrograph unless otherwise stated
any one from:
idea that can see internal structures ;
A example of named cell structure
cannot see surface contours / AW ;
A not 3-D appearance
AVP ; e.g. ref. to small(er) depth of field

c) Calculate the actual diameter, X–Y, of the mitochondrion labelled in Fig. 1.1.
Write down the formula that you will use to make your calculation. Give your answer to the nearest whole nanometre (nm). [2]
Formula:
Actual diameter: correctly stated formula ; e.g. actual diameter = image length / magnification
correct actual diameter of X–Y calculated from measurement from printed copy ;
A 213 nm (for 10 mm)
A 223 nm (for 10.5 mm)
A 234 nm (for 11 mm)
A 245 nm (for 11.5 mm)
A 255 nm (for 12 mm)
SQ 3