Rough Measurements With the Light Microscope
A light microscope is an instrument that allows you to observe specimens to determine what is
present in a sample and what it looks like. However, it is also possible, and generally very
useful, to use a light microscope to measure the size of a specimen under view.
To make these measurements you have know the diameter of the field of view. The field of
view is whatever you can see in the microscope at any given moment and it is smaller at higher
magnifying power.
To measure the field of view you would generally use a very small insert in nthe ocular that is
marked off at regular intervals. The insert is called an ocular micrometer but we will be using a
much simpler devise to measure the field of view-a plastic ruler! By placing the ruler on the
microscope stage at low power you will be able to measure the field of view directly in
millimeters.
Use the following procedure.
Put the clear plastic ruler on the microscope stage with the scanning objective in the viewing
position. You may have to manipulate the ruler to get it to fit on the stage. Put the lines and
marks of the ruler up.
Focus in on the markings of the ruler. The depth of field of the scanning objective should make
this east.
Use your mechanical stage to move the markings so that they stretch across a full field of view
diameter and are aligned with the edge of the field.
When you have focused and lined up the ruler as described above you will be able to read off
the diameter in millimeters. Write down your data! This is the diameter of the field of view with
YOUR SCANNING OBJECTIVE! What is the magnifying power of your objective? Record that
piece of data as well.
For higher power objectives you will probably not be able to use the ruler directly. Therefore
use the following formula to calculate field of view diameter for the low power objective.
(FOV-4X)*(Mag-4X) = (FOV-10X)*(Mag10X)
Where
(FOV-4X) = diameter of field at 4X (what you just measured)
(Mag-4X) = magnifying power of 4X objective (ie 4)
(FOV-10X) = diameter of field of view at 10X (what you want to find out)
(Mag10X) = magnifying power of 10X objective (ie 10)
Substitute the known quantities into the equation, rearrange and solve for the new field of
view.
You can plug in the numbers for your high-dry objective and calculate the field of view for it as
well.
Suppose the field of view diameter is 12 mm with the scanning objective and you can see a long
plant cell in the field of view that stretches one-fourth of the way across the field. Thus you
know that the plant cell is one-fourth of 12 mm in length (1/4 x 12 mm = 3 mm). The plant cell
is 3 mm in length.
Using millimeter rulers makes for east measurement of the field of view diameter but
millimeters are not good units for microscopy. They are too big. Therefore scientists typically
use a smaller unit called the micrometer (u or um) for expressing the size of microscopic
objects.
One millimeter equals 1000 micrometers. So the hypothetical giant plant cell above is 3000 um
in length.
Use the techniques described above to observe some microbes in pond water and to measure
their size.
Protists will be 200-300 um in length at the biggest. Anything larger than this is an anaimal or
plant.
Bacteria are less than 10 um in length so you may not be able to see any of them.