Cambridge International A Level Biology Revision Guide

Chapter P1: Practical skills for AS
BOX P1.2: Using a grid or haemocytometer to count cells
You may need to count the number of cells in a given area
or volume of a sample. To count the number in a given area,
you could use a grid in the eyepiece of your microscope or
marked on a special slide. You can then count the number of
cells in a known area of the grid.
To count the number of cells in a known volume of
liquid, you can use a haemocytometer. You could use it, for
example, to count the number of red blood cells in a known
volume of a blood sample, or the number of yeast cells in a
known volume of a sample from a culture.
The haemocytometer is a special microscope slide with
two sets of ruled grids in the centre, with deep grooves
either side. Between the grooves, the surface of the slide
is 0.1 mm lower than elsewhere. This produces a platform,
called the counting chamber (Figure P1.5).
IMPROVED
NEUBAUER
Depth 0.1 mm
1/400 mm 2
slide while
pressing on edges
coverslip
Figure P1.6 Preparing the haemocytometer.
Newton’s rings
are seen here
Now use a pipette to carefully introduce your sample into
the space between the coverslip and the platform. The liquid
should cover the platform completely, but not spill over into
the grooves (Figure P1.7).
counting positions of
chamber ruled grids
IMPROVED
NEUBAUER
Depth 0.1 mm
1/400 mm 2
IMPROVED
NEUBAUER
side view
top view
251
Depth 0.1 mm
1/400 mm 2
Figure P1.5 Haemocytometer.
To use the haemocytometer, a special coverslip is placed
over the counting chamber. It must be slid into place and
pressed down firmly until you can see ‘interference’ colours
where the coverslip overlaps the slide. (These are called
Newton’s rings.) When you can see these, the distance
between the rulings on the slide and the coverslip is exactly
0.1 mm (Figure P1.6).
Figure P1.7 Haemocytometer containing the sample.
Place the haemocytometer on a microscope stage, and
use the low-power objective to locate one of the ruled
grids. Move up to the ×40 objective, and focus on the block
of squares at the centre of the ruled grids (containing the
numbers 1 to 5 in Figure P1.8). This is made up of 25 squares,
each containing 16 smaller squares. Count the number of
cells you can see in any five blocks of 16 small squares. If
a cell is on the boundary line at the top or on the left, it is
counted. If it is on the boundary line at the bottom or right, it
is not counted (Figure P1.8).
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