BIO202, Human Anatomy and Physiology Name:____________________

1) Microscopy and Cellular Structure Handout Section:___________________

For today’s exercise you will need to

become practiced with the laboratory
microscope. You will be using this
instrument nearly every week for the entire
two semester sequence of Human Anatomy
and Physiology. Becoming familiar with how
to use this tool is an essential skill for you to
develop. Think of the microscope as a tool
for visualizing the form of things at a level of
detail your unassisted eyes cannot see.
Typically, this is done by placing these
viewed objects on glass slides and projecting
light through them. Except for in today’s lab,
you will always be studying biological
tissues. Keep in mind these tissues have
been sliced very thinly (<1-2 cells thick) and
then stained with various chemical
combinations. When you view these tissues,
it is easy to appreciate that the sample has
a surface area but it’s less obvious to the
beginner that it also has depth. This means
with a little practice at the controls, and
with experience learning basic histology,
you can see things under the microscope and appreciate it in three-dimensional space. With this
instrument you control the illumination (e.g., condenser, iris diaphragm, light source) and the
surface height that you want to see in clear detail (e.g., diopter adjustment, ocular lens, course
focus, and fine focus).

Please Read These Important Reminders

A. Always begin with the revolving nosepiece set at the lowest magnification (4x objective).
B. Set interpupillary distance as if you were trying to look through a pair of binoculars. The
ocular lens should be correctly focused. Both of these will be likely be different for each
person in the lab group.
C. Use the 2-way stage control knobs (grips on the right side) to find your slide specimen in
the vertical and horizontal axes. Focus with the course adjustment ONLY ON THE LOWEST
MAGNIFICATION (4x). Use the fine adjustment at higher levels. You do not need to adjust
the stage when you are switching to a higher magnification power. IF YOU ARE IN THE
CORRECT FOCAL PLANE, ALL LENSES WILL PASS ABOVE THE SLIDE WITHOUT CRUNCHING
INTO IT.
D. When satisfied, reset the revolving nosepiece to the 4x objective for your next slide.

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To begin you will need to pull the slide that depicts a letter “e” (slide #95). First make sure you
are viewing it correctly (i.e., it is oriented properly to your eye). Next place it on the stage using
the stage clamp and turn on the light source. With the 2-way stage control knobs place the
specimen in the path of the light and begin to focus.

A. B. Review the two diagrams to the left. They

are meant to depict how light travels to and
from your eye as you view something with a
simple microscope (A) and a compound
microscope (B).

1)What is the primary difference between

the two?
___________________________________

2)Study the original object (small arrow) and

compare it to the resultant image (big
arrow). What are two differences found
between the resultant images in panel B vs.
panel A?
___________________________________

___________________________________

Now confirm your answer to number 2) by studying the letter “e” on the slide with, and without,
the 4x objective. Draw a representation of your field of view at 4x magnification. Try to draw the
letter at the appropriate size relative to the field of view boundaries.

3)What has happened to your letter?

_________________________________________

4)What does this mean for using the 2-way control

knobs if you want to move the field of view up,
down, left, or right?
_________________________________________

_________________________________________

_________________________________________

_________________________________________

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 In the next activity you will need the slide of a grid
with a square size of 2.5mm (slide #1). This is like
having a ruler where each line (side length of the
square) is a known length. Thus, it can be used to
infer the width of your field of view at each
magnification. See the image to the left as an
example of a square grid being used to measure
across a field of view.

5)Why is it important what portion of the field of

view you decide to lay your line of squares across (i.e., if you measure across the middle vs. along
an edge of the circular field of view)?

______________________________________________________________________________

Because this is a compound microscope you use an ocular lens and multiply its magnification with
one of three objective lenses. Confirm that the ocular lens is 10x and the three objective lenses
you will use are 4x (scanning), 10x (low power), 40x (high power), 100x (oil immersion).

6) What is the total magnification by using each objective lens “compounded” by the ocular lens?
Objective lens magnification Ocular lens magnification Total magnification
4x X 10x = ____________
10x X 10x = ____________
40x X 10x = ____________
100x X 10x = ____________

7) Next, you will need to estimate the field of view size in mm for each of the four objective lenses
(4x, 10x, 40x, and 100x). Use the grid slide as a ruler to measure how large the size is of the
scanning lens field of view.

Scanning lens field of view = 2.5 mm per line segment X the number of square line segments
you can count under the scanning lens. Solve for this simple product and insert it below for the
scanning lens field of view.

Scanning power lens @ 4x magnification = _________ mm field of view diameter

Now you will need to use this diameter that you measured to solve for the other three objective
lens fields of view. The reason you need to solve for them is because your grid slide is no longer
an effective ruler to measure with at these greater magnifications. You will need to use this
formula. The “unknown” term refers to the one of the three lenses you are solving for.

𝑆𝑐𝑎𝑛𝑛𝑖𝑛𝑔 𝑙𝑒𝑛𝑠 𝑑𝑖𝑎𝑚𝑒𝑡𝑒𝑟 × 𝑆𝑐𝑎𝑛𝑛𝑖𝑛𝑔 𝑙𝑒𝑛𝑠 𝑚𝑎𝑔𝑛𝑖𝑓𝑖𝑐𝑎𝑡𝑖𝑜𝑛

"𝑈𝑛𝑘𝑛𝑜𝑤𝑛" 𝑑𝑖𝑎𝑚𝑒𝑡𝑒𝑟 =
"𝑈𝑛𝑘𝑛𝑜𝑤𝑛" 𝑙𝑒𝑛𝑠 𝑚𝑎𝑔𝑛𝑖𝑓𝑖𝑐𝑎𝑡𝑖𝑜𝑛

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Low power lens @ 10x magnification = _________ mm field of view diameter

High power lens @ 40x magnification = _________ mm field of view diameter

Oil immersion lens @ 100x magnification = _________ mm field of view diameter

8) You will use these estimations to further predict how many cells wide the field of view is with
each objective lens. If a typical cell is around 15 μm across (which is equal to 0.015 mm), how
many cells wide is the field of view at high power? Just divide the field of view you estimated
from the scanning power, low power, high power, oil immersion objectives.

Scanning lens field of view = __________mm / 0.015 mm = __________ cells wide

Low power lens field of view = __________mm / 0.015 mm = __________ cells wide

High power lens field of view = __________mm / 0.015 mm = __________ cells wide

Oil immersion lens field of view = __________mm / 0.015 mm = __________ cells wide

In the next activity you will use the slide that displays a triad of silk threads (slide #96) to
appreciate the depth of viewing at each level of magnification. Inspect the threads with the
scanning objective and make a guess as to which thread is on the bottom, which is in the middle,
and which is on top. View it at each successive level up to high power. Use the fine focus to test
your prediction. It will help if you use the iris diaphragm to reduce the light coming through.

9)What was the stacking order and how did you know this?

______________________________________________________________________________