DEPARTMENT

OF BIOTECHNOLOGY
KULLIYYAH OF SCIENCE



LABORATORY MANUAL
CELL BIOLOGY
(BSBT 1233)



Module:
Cells and Cell Cycle



Semester 2, 2022/2023

Instructor:
Dr. Nur Firdaus Isa (Coordinator)
Dr. Nur Hafizah Azizan

Lab Personnel:

 TABLE OF CONTENT

Laboratory Operations and Safety

Lab 1: Virtual microscopy and pre-lab exercises

Lab 2: Seeing cells

Lab 3: Mitotic Index

Lab 4: Mitotic inhibitory and level of genotoxicity

The laboratory manual is only a guide. Some changes may occur during the lab
sessions, and these must be documented and described in the lab report.

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LABORATORY OPERATIONS AND SAFETY

Safety is an important consideration in any laboratory environment. The following sections
list safety rules appropriate for any laboratory. Review and familiarize yourself with these
procedures so that your laboratory experience will be an enjoyable and safe one.

GENERAL LABORATORY OPERATING PROCEDURES

1. No food or beverage is to be taken into a laboratory where accidental hand-to-mouth
contamination or ingestion can occur. This is one of the most common ways in which
dangerous microbes can enter the body.
2. Never place personal items such as bag, purse, mobile phone, or clothing on your lab
bench. These items are likely to get stained, become dirty, and contaminated by
microbes. Leave your bag outside the lab.
3. Wearing a lab coat is compulsory. Lab coat protects you from various stains,
chemicals, and microbes (including those in aerosols). Store the lab coat in a sealed
plastic bag before leaving the lab. Lab coat is not for street use!
4. Wear proper covered shoes. Sandals or open-toed shoes are not allowed in the lab.
5. Gloves are provided for protection. Avoid touching your face or rubbing your eyes
while wearing contaminated gloves. Also avoid touching doorknob while wearing
gloves to prevent cross-contamination.
6. Be aware of the locations of the first aid kits, fire extinguisher, eyewash station, deluge
shower, and exits. It is too late to study a floor plan in case of emergency.
7. Follow your instructor’s instructions. Never work in the lab unsupervised, perform
your own experiments without given permission, or engage in childish antics. When
in doubt, ask your instructor or demonstrator.
8. Never put anything in your mouth while in the lab. No solutions should be pipetted
by mouth.
9. Be cautions with chemicals. Never use a substance or chemical that is missing a label.
Always label your samples.
10. Never pour chemicals down the drain without permission, leave any heated materials
unattended, or place flammable substances near heat.
11. Inform your instructor about any accidents, spills, or potential hazards.
12. Discard used or broken slides in the designated sharps containers or disinfectant
solution. Do not handle broken glass with bare hands.
13. Wash your hands prior to and after completing the lab session. Do not bring anything
up to your mouth or eyes without washing your hands first.
14. Before leaving the lab, replace your lab stool or chair under the table. Wipe and clean
your work area with disinfectant. Make sure all materials have been returned to their
original locations.
15. Initial the attendance form after the lab practical session ends.

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 READ THE FOLLOWING INFORMATION CAREFULLY.


Be punctual
Punctuality is a key to academic success. Principle cell biology lab requires you to master
various techniques and needs you to handle all specimens / samples safely and efficiently.
Instruction and demonstrations of these procedures will be done at the beginning of each
session. Any additional information regarding the class or modifications of protocol will
also be announced at the beginning of the session. It is in your best interest not to be late!

Be prepared
If instructed to do so, read the introductory material ahead of time so that you will know
what to expect and what is expected of you. Make sure you have your laboratory manual,
lab coat, marking pen, and whatever else is required for each session.

Be responsible
Take care of your work area and equipment assigned to you. Wipe down your work area
with disinfectant solution before and after each laboratory session. Keep your microscope
clean and in good working order. Follow your lab instructor’s direction for cleaning, setting
up, and putting away your microscope. Be aware of the proper containers needed to place
used slides, tubes, stains, chemicals, paper, and other items used in the lab. Leave the
laboratory area in good order and return all materials and equipment to their original
location.


APPENDICES
SUPPLEMENTARY MATERIAL: USE OF THE LIGHT MICROSCOPE

The microscopes that you will be using are delicate, expensive instruments, and to get the
best results they must be used with care and skill. Please read these instructions carefully
and follow them.

1. Switching on

This procedure must be followed in order to ensure that the bulb does not blow:
i. Check that the rheostate voltage control slide (A, on the right hand side) is in the
position closest to the front. This is the lowest power.

ii. Only now use the on/off switch (B) at the right side to turn on the power.

iii. Power can be increased by sliding the rheostat away from you.

2. Adjusting the eyepieces

The distance by which the eyepieces are separated is adjusted simply by pushing or
pulling apart the sliding base plates. Change them to fit the design of your head, and
thereby minimize eye fatigue. A scale above the right hand eyepiece shows the setting.
Move them whilst looking down both simultaneously until the two images coincide.
Both eyepieces (H) are focusing eyepieces. Set the right hand eyepiece at the middle of
the range position and then, without readjusting that setting:
i. Close your left eye whilst looking down the right eyepiece with your right eye.

ii. Focus the specimen sharply using the focusing knob (M).

iii. Close the right eye whilst observing the specimen down the left eyepiece with
the left eye.

iv. If the specimen is not sharp, focus it by using the knurled ring around the left
eyepiece.

v. When both eyes are open the images given by both eyepieces should now
coincide and be in focus.

3. Focusing on the specimen

Care must be taken while focusing not to damage the lenses. This is especially
important whenever using the higher power (40x and 100x) objectives.
i. Swing the lowest power objective into place.
 ii. Place the specimen to be examined on the stage. Centre the specimen using the
stage movement controls.

iii. Lower the stage using the coarse focus knob (M) WHILE LOOKING AT THE
OBJECTIVE FROM THE SIDE. Be certain that the stage is going down and not up
or your objective will crash through and break the slide. This is likely to damage
the objective.

iv. Look through the eyepieces and focus on the specimen by slowly racking up
the stage (knob M) until the image comes into focus. Note that the focusing knob
consists of an outer ring, the coarse adjustment, and an inner ring, the fine
adjustment. Adjust the focus with the coarse adjustment first and then the fine
adjustment.

v. To examine the specimen at higher magnification, rotate the next objective lens
into place while looking from the side. Make sure the lens does not touch the
slide. Do not adjust the focus.

vi. Examine the specimen through the eyepieces. Minor focusing with the fine
adjustment may be required to bring the specimen into focus. These
microscopes are PARFOCAL, meaning that the higher magnifications should be
in focus once the specimen has been focused on correctly at the low
magnification.

4. Focusing the condenser

It is important to focus the condenser each tie that you start to use the microscope. Only
when it is correctly adjusted will the instrument perform at its optimum and resolve
detail most efficiently. Correct adjustment is also the least tiring on your eyes.
i. Place a prepared slide on the stage. Focus on the specimen using knob M and
with the low power objective in position.

ii. Close the condenser iris diaphragm by swinging lever G away from you. In this
position the specimen on the slide will have a minimum amount of light.

iii. Focus the condenser using the knurled knob E on the left side of the microscope.
First rack the condenser up until it ALMOST touches the slide. Now look through
the eye pieces and rack the condenser down (knob E, turn towards you) until
there is a “grainy” appearance, then focus upwards slightly (knob E) until this
appearance is lost.

5. Open the iris diaphragm

The iris diaphragm is adjusted by a small lever (G) emerging from the condenser
housing. Open the iris diaphragm by swinging lever G towards you as far as it will go.
This is NOT USED TO CONTROL THE INTENSITY OF THE LIGHT REACHING THE
EYEPIECES. Only the rheostat control should be used for that task. If the setting is too

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 narrow there will be interference lines around the edges of the specimen and resolution
will be compromised.
For this reason the iris diaphragm should be left between 100 and 90% open. Closing
the diaphragm will increase contrast. This helps with low contrast specimens but
resolution will be impaired.
Useful information about the objectives on your microscope
Objective Total NA Resolving Use oil
magnifying magnifying power
power power (10x eye
piece)
4x 40x 0.1 3.23 mm NO

10x 100x 0.25 1.29 mm NO

40x 400x 0.65 0.50 mm NO

100x 1000x 1.25 0.26 mm YES**

** The black ring around the lower part of the 100x objective is the universal
microscopy symbol that indicates that the lens should be used with immersion oil.
Only use immersion oil on this objective lens. If you accidentally get oil on the other lens
get a demonstrator to help you clean it.

6. Locking the devices

There are four locking devices for locking various parts of the microscope (I-L). Do NOT
touch any of these. If you feel that one is not adjusted correctly ask a demonstrator.

7. Cleaning lenses

Obviously it is important to have clean lens. Be very careful how you clean these – the
lenses cost up to RM1500 each. ALWAYS USE LENS TISSUE WHEN CLEANING LENSES.
DO NOT REMOVE EYEPIECES, LENSES OR CONDENSER AND ATTEMPT TO CLEAN
INSIDE. ONLY CLEAN THE OUTSIDE OF EYEPIECES AND LENSES USING LENS
CLEANING FLUID AND LENS TISSUE – DO NOT USE COARSE PAPER TOWELS.

8. Putting away

i. Turn the objectives so that the lowest power objective is in viewing position.

ii. Slide the rheostat voltage control slide (A) to the lowest setting (i.e., closest to
the front).

iii. Switch the power off (switch B).

iv. Replace the cover and put away under the bench. Fold the cord neatly beside the
microscope.

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9. Carrying

Always carry the microscope by its arm and be ready to support the base in case this is
loose. Carry it vertically so that the ocular, mirror and filter won’t drop out.