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BIOLOGY 1

HOW TO USE
MICROMETER ON
C

MICROSCOPES
FIS 1707-8019
INTRODUCTION
Most of the time, we would use a ruler to measure objects, but what about
objects that cannot be seen with the naked eye? Of course, microscope is needed in
order to view and measure these small objects. When using a microscope, generally
magnification is higher and a normal ruler will not work. Ocular micrometer and stage
micrometer plays an important role in measuring objects using the light microscope. An
ocular micrometer is used to measure the real size of objects, and stage microscope is
used for calibrating the ocular micrometer. When you look through the eyepiece of a
microscope that has a reticle installed in it, the ruler is super-imposed upon your image.
This makes it easy to make quick and accurate measurements, as long as you have
calibrated your microscope.
MEASURING DEFINITIONS
Stage Micrometer: a microscope slide with a finely divided scale marked on the
surface. The scale is of a known true length and is used for the calibration of optical
systems with eyepiece graticule patterns.
Ocular Micrometer / Eyepiece Reticle:
a glass disk that fits in a
microscope eyepiece that has a ruled scale, which is used to measure the size of
magnified objects.
Field of View: the diameter of the circle of view when looking through the microscope.
CALIBRATION OF THE MICROSCOPE
THE IMPORTANCE OF CALIBRATING THE MICROMETER OF THE MICROSCOPE:
• To ensure the microscope will output accurate and valid information

HOW TO CALIBRATE THE MICROSCOPE:


1. Align the zero line (beginning) of the stage micrometer with the zero line (beginning) of the
reticle
2. Carefully scan over until the lines finally line up
3. Use a simple ratio to determine the value that each line represents in the reticle
4. Recalibrate is a must after switching to a different objective

The larger the number on the eyepiece micrometer, the more accurate your calibration will
be.
IN ORDER TO DETERMINE THE DISTANCE BETWEEN THE LINES ON THE RETICLE, THIS FORMULA
IS NEEDED FOR CALCULATION:

Reticle Division / Objective Lens Value = Distance between lines on reticle at this
objective value
If you are using a 5mm reticle with 100 divisions, your reticle division is 5/100 = 0.05mm
If you want to determine the distance between lines using the 4x objective the formula is:
0.05mm / 4 = 0.0125mm when using the 4x objective.
ACTIVITY 1: BASIC MICROSCOPY
Determine the size of the field of view for each of the other objective lens on your microscope
by using ocular and stage micrometer:
1. Rotate the ocular until the line of the ocular micrometer parallel those of the stage
micrometer by using objective lens 4X, 40X and 100X
2. Align lines at the left edges (0 lines) of the two micrometers by moving the stage
micrometer
3. Calculate the distance in millimeters between lines of the ocular micrometer.
y ocular spaces = x stage spaces
y = 10
x=7
Since the smallest space on a stage micrometer = 0.01 mm, then
y ocular spaces (mm) = x stage spaces X 0.01
1 ocular space (mm) = (x/y) X 0.01
Therefore, if a specimen spans SEVEN spaces the ocular micrometer with that objective in
place, the specimen is 49 μm.