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Laboratory 1
Introduction to the Botany Lab
Microscopes
I. Introduction to the Botany Lab
A. Lab Organization
The botany lab has a basic organization that makes materials, demonstration material, and reference
pictures and charts easy to locate.
1. Typical Student Station
compound microscope (left cabinet)
full set of prepared slides (black slide box)
stereo dissection microscope (right cabinet, but only at some stations)
2. Each End of Each Student Bench
glass slides
cover slips
distilled water bottle
Gram’s iodine stain bottle
neutral red stain bottle
dissection needles
forceps
hand lens
lens tissue packet
plant materials, as needed
3. Perimeter Benches (in order of lab coverage)
plant and other materials
workstations
demonstrations, charts and illustrations
4. Bulletin boards and hanging charts
additional support material
Warning: Most labs are accompanied by a wealth of support material.
Use:
 your instructor’s advice
 the lab manual
 your own interests
to pick and choose the most important components. Your success in the course can be very much
influenced by your ability to use the labs with discrimination.

B. Housekeeping
You share your workstation with students from other sections, so please keep it in good condition. Please
restore every place you work to a condition as good as, or better than, you found it. Let the instructor
know if there are problems passed on to you from previous users.
Each station is equipped with a box containing the entire semester's set of prepared slides. This is a
great convenience, but requires a little collective maintenance. Please do your best to keep slides clean,
in order, and in place according to the key on the box lid. Try to avoid mixing slides between sets. Make
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the instructor aware of missing or damaged slides, because you are liable for slide breakage (see Biology
Department policy statement below).
Slide use guidelines:
(1) Since a complete box of slides represents a large investment, handle it carefully and keep it away
from edges or corners of the lab bench where it could be inadvertently knocked off.
(2) Use one slide at a time unless an exercise specifically requires more than one. Replace each
slide in the box in the right place when you are done with it and before you take another. Never set
slides on papers, books or tabletop. That often leads to accidental breakage.
(3) Pay attention to how you put a slide into the mechanical stage. Keep stage scrupulously clean
and dry. Carefully clean all traces of immersion oil from sides and stage.
(4) When you finish with slides that you have made yourself, dispose of the plant material in an
appropriate way (bacterial slides get dipped in disinfectant). Then wash the slides at the sink, dry
them, and return them to the slide boxes. Throw away scratched cover slips, but please clean and
reuse ones that are still in good condition.

C. Biology Department Equipment Policy

(1) Students will not be charged for class use of expendable materials such as chemicals, seeds,
and cultures.
(2) Breakage of equipment (such as glassware or slides) during class use will not result in a charge
if:
(a) the student returns the broken or defective equipment to the instructor or the stockroom,
and
(b) the student has followed proper laboratory techniques. (See limitation in part 3.)
(3) Breakage will result in a charge if:
(a) the broken or defective equipment is not returned to the instructor or stockroom, and/or:
(b) the individual student's breakage total exceeds that expected despite using proper
technique.
(i) “Normal” is considered to be breakage of not more than three items per semester
or not exceeding a total value of $10.
(ii) Microscope slides fall into the “normal breakage” category for the first four weeks
of the semester only. After that, we assume that any student will be able to use
proper microscope technique at all times and simply not break slides. Any
prepared slide damage that occurs after week four will result in a charge equal to
the cost of replacing the slide.

D. Botany Open Study Laboratories

Normally, both botany laboratories are open two evenings per week for continued study or review. The
laboratories contain the same materials that were present during the scheduled labs that day, so there is
typically a selection of material from two different course units. Since students from all sections share the
study labs, there is no expectation or guarantee that students will work at their assigned lab stations.
For the sake of all users, students attending the study labs should observe the following guidelines:
(1) Use the laboratories for study and review only and go elsewhere for socialization or interaction
that would be disruptive to academic users.
(2) Follow department breakage guidelines (see above).
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(3) Leave materials and equipment where they belong. This includes:
(a) Removing no laboratory materials from either laboratory, even temporarily;
(b) exchanging no materials between laboratories. In study labs, students may work at any
vacant lab station.
(4) Follow instructions of the proctor about appropriate laboratory use.
While the proctors can be a great source of help and information, they should not be expected to have all
the answers. In general, you should try to do as much during scheduled labs as possible, and use the
study labs primarily for review and enrichment.

II. Microscopes:
For more than half the course, your compound microscope will be the most important, most used piece
of equipment. The compound microscopes are binocular, for viewing ease, but the image is transmitted
through only a single objective lens, so stereo vision is not possible. The lab is also equipped with
stereoscopic, broad-field microscopes. These allow true three-dimensional viewing as well as direct
manipulation of viewed material, but they don't provide enough magnification to work at the cellular level.
Such microscopes are also called dissecting microscopes.

A. Microscope Use and Care Guidelines:

(1) Form the habit of using both hands to carry the microscopes, one supporting the base, and one
stabilizing the upper part. Our compound microscopes are formed from one integral unit, but many
excellent microscopes have lens column and base held together only by the rack and pinion used in
focusing, as do our broadfield microscopes. Hold the microscopes upright so that the eyepieces do
not fall out. It is possible to lock eyepieces into a microscope tube, but then it is inconvenient to
switch eyepieces.
(2) Keep lenses as clean as possible! A large proportion of alleged microscope malfunctions turn out
to be dirty objective lenses. Note that some of the objective lenses are somewhat recessed, and
therefore a little hard to clean. Clean lenses only with lens paper!
(3) Keep slides and cover slips clean. Starting with a dirty slide or cover slip is likely to be a waste of
your time. Try to recycle (wash and dry) slides and cover slips whenever possible, but discard cover
slips that are dirty or scratched. Be sure to ask the instructor for cover slips or slides if either is
missing.
(4) Move the distance between the eyepieces (interpupular distance) until you comfortably see a
single image. If you wear glasses, experiment with keeping them on or leaving them off. If your
vision needs lots of correction, it will probably work better to get used to wearing your glasses while
using the microscope. Our microscopes are highly adjustable, but if you change the focus of an
eyepiece from standard, please return it to normal before putting the microscope away, for the sake
of the next user.
(5) If only one side of your view is in focus, bring the image into clear focus for your right eye using
the coarse and fine focus knobs. Then adjust the left eyepiece focus by rotating the knurled diopter
adjustment ring on the eyepiece. If you are an organized person, you might want to record the plus
or minus diopter adjustment that is correct for you. Similarly, you could record the correct
interpupular distance from the scale at the top of the compound scope eyepiece housing.
(6) Do your best to keep the objective lens of the compound microscope from touching the cover slip
or the slide itself. The 20X, 40X, and 100X objective lenses on our compound microscopes are
spring-loaded, so neither they nor the slide are likely to be broken by contact, but many very good
microscopes lack this feature. The result of a collision between lens and slide can be the breaking of
a slide that represents hours of work, or altering the alignment of the lens, requiring at least $100 for
repair.
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(7) Form the habit of returning the compound microscope to low power before putting it away. With
our microscopes it is merely a courtesy to the next user, but with many microscopes it helps avoid
expensive lens, slide collisions.
(8) Use the various objective lenses of the compound microscope and the zoom feature of the
stereomicroscopes intelligently. Don't shun the lower powers, for they show a larger field of view than
higher powers. Use lower powers for search, then switch to higher powers as needed for detail. If
your microscope is working correctly, only fine focus adjustments should be necessary after changing
objective lenses. In this and all microscope operations, please let the instructor know about
malfunctions.
(9) Match light intensity to viewing needs. Both too little and too much light can compromise
resolution. In general, higher magnifications require more light. Detail can often be enhanced in
relatively transparent material by reducing the light.
(10)With the compound microscope, learn the distinction between controlling light with the
diaphragm and controlling it with the intensity control on the side of the microscope base: When the
diaphragm is mostly closed, more of the total depth of your "thin section" will be in focus, but you will
need a higher lamp intensity. Using high bulb intensities for everything, however, will needlessly
shorten the life of our very expensive microscope lamps. The price of increasing light by opening the
diaphragm is a progressive reduction in the total depth that is in focus. This can actually be useful if
you are trying to infer the three-dimensional shape of an object, or tell if one object is above or below
another.
(11)Before turning off the compound microscope, cool the light source by turning to the lowest
possible setting for about 5 minutes.
(12)Avoid sliding the microscopes across the lab benches, for the vibration can weaken the lamp
filaments and shorten the bulb life.
Let your instructor know of any microscope problems. Effective use of the microscope is important for
much of this course.

B. Parts of the Microscope

1. Compound Microscope
Table 1-1. Compound Microscope Parts Checklist. (See Figure 1-1.)
eyepiece housing body
eyepieces (ocular lenses): 10X coarse focus knobs (large, inner)
interpupular distance slide fine focus knobs (small, outer)
eyepiece pointer (in right eyepiece) pre-focusing lever
eyepiece micrometer (in left eyepiece)
eyepiece focus knob (on left eyepiece) stage
stage slide clip
tube (lenses inside) in/out stage movement knob (upper)
tube swivel lock screw left/right stage movement knob
aperture iris diaphragm, with lever
turret (revolving nosepiece)
objective lenses: 4X, 10X, 20X, 40X, [100X*] base
lamp (inside housing)
condenser lamp intensity adjustment
condenser height adjustment knob power switch
* Will be swapped for the 4X lens when especially high resolution is required.
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Figure 1-1. Compound Microscope.

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Figure 1-2. Broadfield Stereo Microscope.

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2. Broadfield Stereo Microscope

Table 1-2. Broadfield Stereo Microscope Parts Checklist. (See Figure 1-2.)
eyepiece housing tube yoke
eyepieces (ocular lenses): 10X tube yoke lock screw
interpupular distance slide upper lamp
eyepiece focus knob (on left eyepiece) upper lamp housing

tube (zoom lenses inside) base

zoom control knob: 0.67X – 4X specimen plate
paired (binocular) objective lenses sub-specimen lamp (inside)
sub-specimen mirror
body mirror rotation knob
focus knobs switch for upper lamp (left)
switch for sub-specimen lamp (right)

The rest of this laboratory and part of the next one will concentrate on the compound light microscope, the
simplest, most versatile tool for examination of very small objects. If these exercises seem elementary
because of your past experience, be very sure that you can understand and can perform the procedures
covered. If some seem difficult or confusing, be sure to ask for help!
Now, and in future laboratories, share your knowledge and your questions with one another. Learning is
often most effective when it comes informally from a peer, and there is probably no better way to solidify
your knowledge than teaching what you know to someone else. On the other hand, be sure to do
instrument manipulations yourself. Our labs introduce and reinforce skills that, like driving a car, you
can’t really learn second hand.

C. Use of the Compound Microscope

To become familiar with the basic workings of your compound microscope, examine the colored strings,
graph paper squares, and typed letters that should be on the first prepared slide in your set.
(1) Lower the stage with the coarse focus knob.
(2) Be sure the 4X objective lens is in place, rotating the turret if necessary to bring that lens into
position.
(3) Clip the prepared slide onto the stage.
(4) Raise the stage as far as possible.
(5) Use the left/right and backward/forward stage adjustment knobs to center the examination
material under the objective.
(6) While looking through the microscope, lower the stage with the coarse focus knob until material
on the slide comes generally into focus.
(7) Focus on the letters that are part of the slide, and compare their orientation in the field of view
with their actual orientation on the slide.
(8) Use the stage adjustment knobs to move the slide right and left, and forward and backward, and
note how the image movement compares with the slide movement. It is possible, of course, to
produce lens systems that show images that match the material in orientation and direction of
movement, but getting used to the inversion is so easy that few will pay the increased expense.
(9) Rotate the turret to bring successively higher-powered objectives into position, and note the
changes in magnification, clearness of image, and number of letters that you can see. At higher
powers, practice adjusting the fine focus to increase visible detail. Look for inconsistencies in focus in
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different parts of the field of view, especially when using the higher-powered objectives. A research
microscope would have higher quality lenses, and come closer to consistent focus across the entire
field. People who use microscopes a lot constantly adjust the fine focus as they work. Changes in
appearance of an object as one focuses up and down give clues to three-dimensional shape. Later
exercises in this lab will offer increased opportunity to demonstrate that for yourself.
(10)Use the graph paper to note changes in size of field with changes in magnification. What is the
maximum number of entire 1-mm squares that you can see at each power? Complete Q1 on the
answer sheet. Note that the total magnification is determined by multiplying the power of the
objective by the power of the eyepiece (10X).
(11)Examine the intersections of the colored strings using high power (40X objective) and with the
diaphragm opened wide (and the lamp brightness adjusted to a comfortable level). This minimizes
the depth of field (the depth of the region that is in focus). Focusing up and down, determine the
order in which the strings occur from top to bottom. Fill in Q2 on the answer sheet.

Key Words
compound microscope
stage
binocular
meter
objective lens
centimeter
broadfield microscope
millimeter
stereoscopic microscope
micrometer
dissecting microscope
nanometer
eyepiece
depth of field
interpupular distance
coarse focus knob
fine focus knob
diaphragm
ocular lens
turret
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Answer Sheet, Laboratory 1

Q1. Compound Microscope Magnification.

Objective Total Approximate Number of

Power Power Squares
4X

10X

20X

40X

Q2. Depth Perception with a Compound Microscope.

Color of Top String

Color of Middle String

Color of Bottom String

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