Sumayya Arif

Roll# 19-11120
Fall 2019
BIOT 211
Biosafety and Biosecurity Course
Section A
Instructor Dr. Muhammad Imran

PRACTICAL NOTEBOOK

1
 TABLE OF CONTENTS

Page
Lab No. Date Activity
No.

1 24/09/19 Laboratories’ Tour 3-12

2 01/10/19 Code of Conduct in a laboratory 13-15

3 22/10/19 Instrument-Based Risk Assessment Of A Sonicator 16,17

Reading, downloading, printing, signing, and attaching

4 29/10/19 Material Safety Datasheet (MSDS) of the required 18,19
chemicals with the lab copy

5 05/11/19 Procedural Risk Assessment 20-22

6 09/12/19 Autoclaves’ Presentation 23

September 24th, 2019 LAB # 1

2
ACTIVITY: Laboratories’ Tour

The instructor took the whole class to FCCU’s research-based biolabs, to give us a rundown of all the
laboratories’ design features, construction, containment facilities, equipment, hazardous materials/chemicals,
microbial agents, work practices, and operational procedures. He instructed us to take note of the safety
equipment, containment facilities (2ndry barriers) placed in those labs and to jot down the purpose of each
instrument, the risk those instruments pose, and Biosafety level of each lab.

PURPOSE:
 To know locations of safety equipment and emergency procedures (i.e. laboratory safety showers,
eyewash stations, and fire extinguishers)
 To observe the implementation of the most common General Laboratory Safety Rules being followed
in every laboratory
 To determine the BSL of each one of the visited laboratory

1. MOLECULAR DIAGNOSTIC LAB (Room#443)

PRACTICES
 Neither Biohazard warning sign was displayed nor Biosafety level (BSL) was clearly stated on any label.
 Eatables were found to be there on lab shelves.
 No proper waste disposal (traditional bakery items found in bins). Though biohazard waste bins were
there but biohazard waste disposal bags were not placed in them
 Sink located close to the door but wasn’t not sensor-operated
 Doors partially closed
 open bench work
 Biohazard infectious waste Puncture resistant sharps containers were present (i.e. a 1º physical
containment device for tips, sharp materials, blades etc.)
 Emergency exit route was displayed on the backside of the door.
 BSC was present in a safe and separate compartment but without any UV hazard sign.
This lab has much better physical containment and it is compartmentalized, and with just some improvements
it can be categorized into a standard BIOSAFETY LEVEL 2 LAB.
Equipment Use Hazard
a common piece of Electrical spark hazard (either from the on-off switch or the
laboratory equipment used bimetallic thermostat used to regulate temperature)
for evenly heating samples
(without any exposed Burn hazard from accidental contact with a hot surface
Hot Plate flames) or for
mixing/stirring solutions in Fire/explosion hazard if the device is located too close to
the laboratory flammable or combustible materials
Vortex Mixer a must critical piece of Accidental spill and contamination hazard
(Vortexer) laboratory equipment on any Electrical hazard
lab’s bench used to mix
samples in test tubes, flasks, Falling may occur due to spillage, hazardous chemical
well plates, or small vials reagents could cause discomfort or possible tissue damage
(via agitation) for cell
disruption or Potentially hazardous aerosol release

3
 homogenization
Burn hazard

Spillage hazard from superheated agarose

Accidental exposure to potentially toxic mutagenic chemicals

like Et.Br. through spill, splash

Inhalation of toxic vapors

Horizontal
used for the separation and
(Agarose Gel) Aerosols’ generation hazard while using pipetting
staining of DNA fragments
Electrophoresis
Equipment Electric shock hazard (b/c of high voltage and conductive
fluid, leaking chamber, corroded electrode cables, or faulty
power supply) causing damage to the skin, muscles, and
nerves

Radiation hazard to skin and eyes from exposure to UV light

while using trans-illuminator or severe burns from Heated
Agarose in a Microwave Oven

Occupational illness due to accidental exposure of potentially

hazardous biological agents (e.g. infectious organisms)

Accidental injection, cuts and abrasions

the most effective primary
containment device for
Occupational illness due to accidental exposure of chemical
operations involving
Biological (e.g. carcinogens) and radiological hazards as these are
potential splashes, spills, or
safety Cabinet commonly used in microbial procedures
aerosolization of hazardous
biological materials
(Biocontainment Fire hazard may occur if somehow the vapors from volatile or
It is designed to provide
device) toxic chemicals build up inside the cabinet.
personnel, environmental
and product protection
Spillage or fall hazard due to brakeage of vials or containers
containing cultures contaminated with infectious substances

Radiation hazard due to UV exposure

2. TISSUE CULTURE LABORATORY (Room#444)

PRACTICES
 Neither Biohazard warning sign was displayed nor Biosafety level (BSL) was clearly stated on any
label.
 Doors partially closed
This lab is not spacious at all but do possess several compartments within it, one containing a BSC and
another Climate Tissue Control Room for carrying out plant, protoplast, and tissue culture growth and storage
in a pathogen-free and antiseptic environment. And so with just some improvements it can be categorized
into a standard BIOSAFETY LEVEL 2 lab.
 All of the students were wearing lab coats as per the requirement.
4
 Liquid Nitrogen Dewar was present at an elevated place with its hazard signs.
 The lab was not tidy. Eatables were found to be there on lab shelves.
 No proper waste disposal (traditional bakery items found in bins). Though biohazard waste bins were
there but biohazard waste disposal bags were not placed in them.
 Sink was not sensor-operated
 Open bench work but having coarse shelves with many cracks on them
 Biohazard infectious waste Puncture resistant sharps containers were present.
 Emergency exit route was not displayed on the backside of the door.

5
 Equipment Use
Liquid Nitrogen a specialized type of vacuum flask
Dewar used for storing, pouring,
(Cryogenic pressurized, dispensing, and dipping
Storage Dewar) of cryogens (such as liquid N2, dry ice,
liquid He) and biological sample
Containment preservation
device cryopreservation
a furnace in which the subject
Muffle material is isolated from the fuel and Burn Hazards
furnace/oven all of the products of combustion
(Incinerator) (including gases and flying ash) in
Containment order to determine the percent of ash surrounding environment
device content in that material.
a piece of laboratory equipment used
Shaker to mix, blend, or agitate substances
in a tube or flask by shaking them
a piece of lab equipment i.e. used
Microcentrifug
to spin small (2 ml or less) liquid
e
samples at high speeds
(Microfuge)
It integrates both a shaker and an
incubator into one space-saving unit,
i.e. designed to simultaneously
incubate, shake, or agitate samples
Shaking
(by the distribution of nutrients and
incubator
the fusion of oxygen into a cell
culture) for efficient performance in
applications like cell aeration,
Hazards
Frost Bite (i.e. a serious freezing injury)
Thermal (cold) cryogenic burns due to
extreme cold temperature
Potentially life-threatening asphyxiation
hazard due to oxygen deficiency
(suffocation) if not stored in a well-
ventilated place
Accidental Release or overflow
Potential pollutants’ release into the
Spill being the most prevalent
possible hazard here
Breakage hazard of culture vessels
(resulting in broken glass)
Exposure to pathogens
Breakage of tube being the most
common hazard
Exposure hazards due to aerosolization
of biohazardous, chemical, or radioactive
materials
Physical (Mechanical) hazard due to
Rotor failure or detachment b/c of
imbalanced and mishooked buckets,
corrosion of the rotor over time,
mechanical stress, metal fatigue, or
dropping etc.
Electric Shock hazard
Fire/Explosion hazard (if the centrifuge is
unbalanced)
Potential spill hazard
6
Electric shock hazard
 3. MOLECULAR BIOLOGY LAB (Room#445)
(BSL-2)
PRACTICES
 Neither Biohazard warning sign was displayed nor Biosafety level (BSL) was clearly stated on any label.
 Open bench work but the counters & shelves were cluttered with lab supplies. Sonicator was not
contained in this lab.
 Sink was not sensor-operated
 Though biohazard waste bins were there but biohazard waste disposal bags were not placed in them.
 4°C cabinet was overfilled with contents and was not properly arranged
 Emergency exit route was not displayed on the backside of the door.

INSTRUMENT USE RISKS

a tissue culture hood is a carefully

enclosed physical containment
device that act as a primary barrier Potential release of toxic fumes (no
Laminar flow to prevent contamination of environmental protection)
hood/cabinet biological samples, and any Contaminated air stream blows direct to
particle sensitive materials against the user’s face (no user protection)
airborne contaminants from
entering the cabinet workspace
used for storage of lab samples
4°C Cabinet Electric shock hazard
and controlled cooling of products

Ideal for grinding, blending, Electricity Hazard

chopping, quick homogenization
Blender Aerosols’ generation hazard
during sample preparation for
microbiological testing Spill hazard

It integrates both a shaker and an

incubator into one space-saving
unit, i.e. designed to
simultaneously incubate, shake, or
agitate samples (by the Potential spill hazard
Shaking
distribution of nutrients and the
incubator Electric shock hazard
fusion of oxygen into a cell
culture) for efficient performance
in applications like cell aeration,
solubility studies, and cell
culturing.

4. MOLECULAR BIOLOGY LAB (Room#429)

7
 BIOSAFETY LEVEL: 2
PRACTICES
Labels of biohazards were there. Shelves were tidy. Sonicator was also contained and earmuffs were also
available in this lab.
EQUIPMENT USE
the most effective primary
containment device
Biological Safety used for safely working with
Cabinet infectious agents/pathogens or
(Containment their toxins(requiring a defined
device) biosafety level) to prevent the
escape of biological aerosols into
the laboratory environment
a motor-driven spinning device
used for separating
particles/components of
cells(such sub cellular
Lab Centrifuge
organelles, DNA, enzymes, or
large molecules proteins) from a
solution or a sample mixture via
sedimentation principle
Sonicator high-frequency sound generator
used to agitate cells, disrupt
bilayer membranes or shear
RISKS
Radiation hazard due to UV exposure
Occupational illness due to accidental
exposure of potentially hazardous
biological agents (e.g. infectious
organisms)
Occupational illness due to accidental
exposure of chemical (e.g. carcinogens)
Fire hazard may occur if the vapors from
volatile or toxic chemicals build up inside
the cabinet
Accidental injection, cuts, and abrasions
Spillage or fall hazard due to brakeage of
objects contaminated with infectious
substances, including vials or containers,
or spilled infectious substances, including
cultures
Breakage of tube being the most
common hazard
Physical hazard of Mechanical failure due
to rotor detachment b/c of imbalanced
and mishooked buckets, corrosion of the
rotor over time, mechanical stress, metal
fatigue, or dropping etc.
Exposure hazards due to aerosolization of
biohazardous, chemical, or radioactive
materials
Electric Shock hazard
Fire/Explosion hazard (if the centrifuge is
unbalance
Hearing damage hazard (caused by high
frequency sound waves)
8
 nucleic acids for the purpose of
cleaning, mixing solutions,
increasing dissolution rate via
mechanical vibrations
Aerosols’ generation hazard from the
sonication process
Chemical burn hazard (as both TEMED &
APS are skin irritants)

Monomeric acrylamide is a potent

neurotoxin

Electric shock hazard (b/c of high voltage

and conductive fluid, leaking chamber,
corroded electrode cables, or faulty
Vertical
power supply) causing damage to the
(Polyacrylamide) commonly used in the lab for skin, muscles, and nerves
Gel the separation of proteins
Electrophoresis based on their molecular weight Accidental exposure to potentially toxic
apparatus mutagenic chemicals

Spillage hazard

Aerosols’ generation hazard while

pipetting

Radiation burn hazard to skin and eyes

from exposure to UV light while using
trans-illuminator

5. METABOLITES LABORATORY (ROOM#428)

(PALEOPALYNOLOGY LAB)
BSL-2

PRACTICES
 Neither Biohazard warning sign was displayed nor Biosafety level (BSL) was clearly stated on any label.
 Sink was not sensor-operated
 Open bench work
 Donned labcoats were not closed properly
 Emergency exit route was not displayed on the backside of the door.
 This lab also has containment and compartmentalization. Centrifuge machine was placed in a
separate compartment in the back so that people might remain safe.
 Punctured Resistant Containers were present. Though biohazard waste bins were there but biohazard
waste disposal bags were not placed in them.
 A separate area for placing bags and personal belongings was available.

Instrument Purpose Risks

9
 Possibility of liquid spill hazard

an enclosed and sterilized vessel Aerosols’ formation hazard

that maintains optimal conditions
for the growth of anaerobic cells Electric shock hazard
Fermenter
and microbes (which undergo
fermentation to produce large Explosion hazard due to accumulation
quantities of a desired metabolite) of carbon dioxide or methane (biogas)

a motor-driven spinning device
used for separating
particles/components of
Centrifuge (with a
cells(such sub cellular organelles,
specific bucket
DNA, enzymes, or large molecules
rotor)
proteins) from a solution or a
sample mixture via sedimentation
principle
Fire hazard
Breakage of tube being the most
common hazard (as sample leakage
cause aerosols, corrosion and
contamination)
Mechanical hazard due to Rotor Failure
or Detachment b/c of imbalanced and
mishooked buckets, corrosion of the
rotor over time, mechanical stress,
metal fatigue, or dropping etc.
Exposure hazard to toxic mutagens

Electric Shock hazard

Fire/Explosion hazard (if the centrifuge

is unbalanced)
Radiation hazard due to possible
a gel image system i.e. widely used
exposure to UV light causing serious
for the imaging
burns to the skin
Gel and documentation of nucleic acid
Documentation and protein suspended within
Exposure to hazardous chemicals that
System polyacrylamide or agarose gels
may be potential carcinogens
(Gel imager) by means of using fluorescence
staining and visualization via
Ethidium bromide is a potential
ultraviolet illumination
mutagen
a type of local ventilation device
i.e. designed to limit exposure to
Mechanical hazard as fume-hood fans
toxic, flammable, noxious, or
are susceptible to failure, thus leaving
hazardous fumes & vapors, gases,
An exhaust fume the hood effectively useless
or dust material by diluting it with
hood
a large amount of air, drawing it
Possible accidental breakage of sash
through an exhaust system, and
cords
then expelling the air in vents
located on the roof of the building.
10
 6. MICROBIOLOGY LAB (Room#427)
(BSL-2)

PRACTICES
 Neither Biohazard warning sign was displayed nor Biosafety level (BSL) was clearly stated on any label.
 Safety signs displayed
 Though biohazard waste bins were there but biohazard waste disposal bags were not
 Sink was not sensor-operated
 Open bench work
 Labcoats were not donned as per the requirement. There were girls who have not tied their hair
properly.
 Emergency exit route was not displayed on the backside of the door
 This lab contains the best BSC available in FCCU’s biology department so far
 Gel documentation system was coded for safety
 Lab supplies and other objects were place at the edges of the shelves which could have fallen quite
easily.
 Surprisingly, this lab has the best containment facilities among the other labs.

11
 Instrument Purpose
an enclosed all-in-one type airtight
centrifugal vacuum chamber used
in proteomics, genomics, cell
biology, microbiology, and drug
development to concentrate
Vacuum Centrifuge/
solutes, DNA, RNA, nucleotides,
Concentrator
and other proteins (from samples
contained in different solvents)
into a dry state or a wet pellet
state by employing Centrifugation
+ Vacuum + Heat altogether
a type of local ventilation device
i.e. designed to limit exposure to
toxic, flammable, noxious, or
hazardous fumes & vapors, gases,
A fume hood or dust material by diluting it with
a large amount of air, drawing it
through an exhaust system, and
then expelling the air in vents
located on the roof of the building.
a gel image system i.e. widely used
for the imaging
and documentation of nucleic acid
Gel documentation and protein suspended within
system polyacrylamide or agarose gels by
means of using fluorescence
staining and visualization via
ultraviolet illumination
an enclosed, ventilated laboratory
Biosafety Cabinet workspace for safely working with
(Class II Type A2) materials contaminated with (or
potentially contaminated with)
(containment device) pathogens requiring a
defined biosafety level)
Risks
Aerosols’ generation hazard
Electric shock hazard
Mechanical failure hazard
Mechanical hazard as fume-hood fans
are susceptible to failure, thus leaving
the hood effectively useless
Possible accidental breakage of sash
cords
Radiation hazard due to possible
exposure to UV light causing serious
burns to the skin
Exposure to hazardous chemicals that
may be potential carcinogens
Gel doc could result in information
leak/misuse if not coded properly
Ethidium bromide is a potential
mutagen
Radiation hazard due to UV exposure
Occupational illness due to accidental
exposure of potentially hazardous
biological agents (e.g. infectious
organisms)
Occupational illness due to accidental
exposure of chemical (e.g.
carcinogens)
Fire hazard may occur if the vapors
from volatile or toxic chemicals build
up inside the cabinet
Accidental injection, cuts, and
abrasions 12
Spillage or fall hazard due to
brakeage of an object
September 24th, 2019 LAB # 2

ACTIVITY: Code of Conduct in a laboratory

Purpose:
As we all know human blunders till date are one of the most crucial factors in turning hazardous incidents up
in the laboratories. Therefore, in order to maintain the highest standard of safety and responsibility we all
need to voluntarily agree to abide by the code of conduct, ethics, and practice providing clear guidelines for
all aspects of appropriate professional laboratory conduct. Everybody should consider these codes and
exhibit effective and safe laboratory behavior before being granted the freedom of working in a laboratory.
Observance of these rules and regulations helps an individual to recognize and resolve the ethics and
compliance issues that may arise in his/her daily work. Deep down, we do not need to be judgmental about
others’ behavior at all as this hinders their learning route, thus making them hesitant in showing a productive
behavior. Rather, we need to first make ourselves able enough than to point others out at their conduct as we
all share the equal responsibility to contribute from the sphere of our professional competence to the
general well-being of society on the whole.

1st Task
The instructor asked us to speak out the very first word that comes to our mind regarding the word
“BIOSAFETY”. All the students participated and the following set of words turned up,
RESPONSIBILITY
taking personal responsibility for the safety, security, and well-being of ourselves, those around us, and our
environment. Holding ourselves accountable for our actions to the institution and the society as we earn the
respect of our community

INTEGRITY
Being honest & sincere, keeping our commitments, and taking liability for our actions and outcomes as this
promotes confidence and trust. Honesty must prevail even if there’s no monitoring

RESPECT
embracing diversity, valuing the perspectives & contributions of others, and acting professionally towards all
by respecting all members of the team

COMMITMENT
dedicating ourselves to helping each other, doing our best

PERSEVERANCE
Staying there in the course until we achieve success.

PLIABILITY (FLEXIBILITY)
being open and accommodating in our actions and decisions

FOCUS
Fulfilling individuals’ needs, creating customized solutions, and providing them a competitive advantage
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MANAGEMENT
Being thrifty and carefully using our resources (entrusted to us)in a safe and responsible manner
COMPASSION
being generous, empathic, considerate, and kind in our interaction with others

TEAMWORK
Working together to achieve common goals and a meaningful purpose in our lives, and the lives of others
APPRECIATION
Valuing engaged and enthusiastic participation by supporting, sharing, and communicating openly &
effectively with each other

EXCELLENCE
exhibiting and exceeding the highest standards of merit in every aspect of our lives

INNOVATION
embracing change by continuously seeking to improve. Thinking creatively, pursuing innovative ideas, and
delivering excellence to positively change our community, nation, and world

SERVICE BEFORE SELF

As everybody matters, everybody counts so without considering self-interests one should enhance lives
through scientific contributions, technology, and good deeds

LEARNING
fostering lifelong learning and striving for excellence in all that we do

2nd Task
The instructor engaged us in an activity of standing in a circle and asked us to count randomly up to the
number 15 without any repetition, observing the following rules,
 Let it be random calling of numbers. “NO PATTERN” should be observed.
 Nobody’s allowed to dictate the other person verbally/symbolically or take lead.
 One should not utter a number until he/she thinks that everyone else has spoken out and it’s his/her
turn again.
 If for any reason more than one person speaks at a time, the counting will restart.
 Everybody has to participate
Although we failed many a times to count till 15 without any repetition but that failure was worth it. It was a
good learning activity which made us realize of what actually was required to complete this task
successfully so that we could stretch out that missing element of learning to the laboratory safety. Infact, this
activity promoted a safe behavioral development in each one of us and made us perceive things positively
even in setbacks. Meaning thereby, if things don't go as planned, one should develop a new strategy to
accomplish his/her task but failure should never be the decision made in response to that particular setback.
The elements of the behavior found lacking in us on the whole were as follows,
 Being proactive
14
 Co-ordination
 Vigilance
 Patience
 Respect
 Time management
 Comprehension (Understanding)
 Even-handed approach (fair and impartial in treatment or judgment)
 Teamwork (mutual accountability & sportsmanship)
 Awareness

15
October 22nd, 2019 LAB # 3

ACTIVITY: INSTRUMENT-BASED RISK ASSESSMENT OF A SONICATOR

Sonicator is a  high-frequency sound generator using high frequency sonic waves to disrupt & lyse cells, to
mix the liquids, or to shear nucleic acids following sonication procedure.

Q1. What can happen?

1. Ultrasonic induced hearing damage (SHRILLING VOICE i.e. a Noise hazard)
2. Breakage of glass container
3. Spillage of microorganism
4. Exposure to chemical
5. Mucosal irritation due to aerosols’ generation (i.e. a Biological hazard)
6. Electric shock Hazard (i.e. use of loose sockets is strictly discouraged)
7. Mechanical failure of the apparatus (such as probe can be broken)
8. Flying object
Q2. Who can be affected?
1. Experimenter
2. Lab personnel
3. Students/fellow workers
4. Lab technician/ Lab attendant
5. Instructor
6. Maintenance (janitorial) staff
7. Repair workers
8. Visitors
Q3. What are the chances of occurrence?
Probability of Occurrence is dependent upon the following,
 Frequency of use of sonicator
 Distance
 Number of people in the lab
 Level of training
 Amplitude
 Quality of wires and connection
 Maintenance level
 Quality of the Container
Q4. How to mitigate?
(Risk Management)
Mitigation strategies are applied to control the identified risk as follows,
1. Guidelines through SOP (no cost)
2. using containment principle (i.e. sonicator located in a transparent sound proof box) 70,000 PKR only
3. Wear earphone-type sound muffs to protect your hearing while sonicating (700 PKR only)
4. Separate room (1 lakh rupees)
5. Moving the sonicator to a less populated area (no cost)
6. Safety glasses (80 PKR only)
7. Putting a limit on the amplitude (as bacterial cells can be lysed at 50% amplitude)

16
 CONTAINMENT TECHNIQUE AT FCCU
At FCCU, one of the Biological Sciences Department’s instructor along with his pupil improvised the
containment of a sonicator by placing that in a wooden box (which in actual was the packaging of a
microscope).
Q5. Who can mitigate?
1. Trainer
2. Instructor
3. Lab incharge
4. DEPARTMENT/Institution
5. University Administration
6. Students
7. Company
The best mitigation strategy against the posed risks can be applied by the ‘Department’, after that by the
University administration, Lab incharge, and lastly by the students.

17
October 29th, 2019 LAB # 4

ACTIVITY: Reading, downloading, printing, signing, and attaching MATERIAL SAFETY DATASHEET (MSDS) of
the required chemicals with the lab copy

• Name of the Material: “Bleach”

Bleach - LabChem Inc. (n.d.). Retrieved from http://www.labchem.com/tools/msds/msds/VT150.pdf.
• Chemical Abstracts Service Registry Number (CAS): 7681-52-9
• Brief Summary of Bleach’s Characteristics & Hazards’:
Bleach appears as a colorless or slightly yellow watery liquid with a characteristic faint chlorine-like odor. It is
a chemical compound comprising up of a sodium cation (Na +) and a hypochlorite anion (ClO⁻) bound together.
Sodium hypochlorite (NaClO-), the active ingredient in chlorine bleach, being corrosive can cause skin burns
and irreversible eye damage, particularly when used in concentrated forms. Therefore, Sodium hypochlorite is
generally used dissolved in water at various concentrations because its dissolution in water forms
hypochlorous acid (HOCl), i.e. a very strong oxidizing agent that can react with and break down many
molecules, including dyes.

Figure of Bleach (Sodium hypochlorite)

NaOCl + H2O → HOCl + NaOH

In laboratories, bleach is used as a sanitizing or disinfecting agent having concentrations ranging from 4 to as
high as 9%. Whereas, domestically it is used as a bleaching agent usually containing 5% sodium hypochlorite
(i.e. a skin irritant with a pH of around 11). Strong bleach use is always discouraged, as it contains a
concentration of 10-15% sodium hypochlorite with a pH of around 13, making it corrosive thus causing severe
skin burns. Sodium hypochlorite in bleach being a strong oxidizer reacts with flammable compounds and
reducing agents causing explosions. Moreover, its solution is a weak base i.e. inflammable. Therefore, users
are recommended to take a number of precautions in order to avoid any personal injury when working with
bleach.
Bleach is used extensively as an oxidizing agent in organic (petroleum) products’ refining, as a bleaching agent
in the textile, detergents, paper, and pulp industries, as a disinfectant in swimming pools, wastewater
treatment, sanitary equipment, and some medical treatments also. Besides this, in food processing, bleach is
used to sanitize food preparation equipment, in fruit and vegetable processing, mushroom production, hog,
beef and poultry production, maple syrup production, and fish processing.
Even though bleach whitens your whites, cleans your floors, and acts as a great sanitizer, it can be quite
hazardous when not handled properly. For instance: its exposure has a pronounced irritant effect on the
skin, its inhalation produces severe bronchial irritation and pulmonary edema, its ingestion causes burning of
mouth, nausea, vomiting, delirium, and coma etc. Bleach should not be mixed with other cleaners, as
reaction with acids can produce toxic chlorine gas.

HClO + HCl → CL2 + H2O

• Name of the Material: “Tween 20”
18
 Home. (n.d.). Retrieved from https://www.fishersci.com/shop/products/tween-20-fisher-bioreagents-
2/bp337100
• Chemical Abstracts Service Registry Number (CAS): 9005-64-5
• Brief Summary of Tween’s Characteristics & Hazards:
Tween is a lemon to amber-colored oily liquid at room temperature with a faint characteristic odor. It is a
polysorbate surfactant with a fatty acid ester moiety and a long polyoxyethylene chain. It is a nonionic bio-
compatible detergent widely used in biochemical applications. Tween 20 is frequently used for lysing
mammalian cells and permeabilizing cell membranes at a concentration of 0.05 to 0.5%. In addition, it is also
used as a solubilizing agent in pre-extraction of membrane-bound proteins and as a washing/blocking agent
in Western blotting (immunoassays) & ELISA to minimize nonspecific binding of antibodies and to remove
unbound moieties. On top of this, it is also used as an emulsifying agent for the preparation of stable oil-in-
water emulsions and as a dispersing agent in pharmaceuticals for placing polyethylene microspheres in
suspension.
Globally Harmonized System (GHS) authenticates tween 20 to be a risk-free, non-toxic substance or mixture
and verifies it to be of low concern. Therefore, it is not expected to be a health hazard even by inhalation.

Figure of Tween 20

November 5th, 2019 LAB # 5

19
ACTIVITY: PROCEDURAL RISK ASSESSMENT

The instructor asked us to take out our cellphones, search an article entitled as “Isolation, Identification, and
Characterization of a New Highly Pathogenic Field Isolate of Mycobacterium avium spp. avium” on Google and
carry out an oral risk assessment of all the procedures used in that article.

Mycobacterium avium
Culture, Staining, Biochemical assays, Genetic analysis
BSL III facility

FOWLS (BIRDS) Specimen: Mass of TB

Gram Staining for gram (+) ve bacteria
Acid-fast staining for acid-fast bacteria (AFB)
Sera samples
Virulent strains Vs Reference strain (as a Control)
Subspecies differ on the basis of Antigenic specificity
Specimen: Tissue containing typical tubercles
Possible Risk Assessment:
• Scratches & bite risk
• Feathers, dust particles from birds’ cage, fecal matter
• Needle sharpened objects’ risk
• Bird's escape risk
Signs and symptoms on the basis of epidemiological information given
Veterinary disease is a one health issue
Being self-proactive matters here
Wearing metal-mesh gloves to circumvent the risks assessed above

NECROPSY
• Aerosols' generation
• Splashes
• Knife cuts and abrasions (as sharp object is being used)
• Contamination of the container containing the specimen
• Splash
• Forceps' mishandling
Now the experimenter is moving towards Diagnostic...
If its dry ice then Frost Bite may occur (i.e. a serious freezing injury)

Thermal (cold) cryogenic burns due to extreme cold temperature

Logistics' mishandling

CELL BLOCKING
Either the chemical properties of such a chemical/material may harm us or that typical chemical/material may
be inappropriate for use
Formalin has been used for fixing of cells and tissue sections while preserving the specimen, and we know that
• Formaldehyde is a potential occupational carcinogen
• Accidental Exposure to formaldehyde can occur which may irritate the skin, throat, lungs, and eyes.

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Anyways, the level of exposure to formalin depends upon the dose, duration, and work being done. So we
need to get to know about a preservative disinfectant’s effect
In vitro diagnostic reagent
Dye used

Ziehl-Neelsen dye (Z-N) technique to detect Mycobacteria

Carbol-Fuchsin Solution for Microscopy
Compound risk assessment is required here
Ziehl-Neelsen is found to be
• Flammable liquid,
• Skin corrosive,
• Mutagen causing germ cell mutagenicity

Z-N Acid Fast Staining

• droplets’ generation
• Splash hazard
• Sharp risk
• Waste generation (contaminated procedure)
• Accidental exposure to Chemical (Saffranin) can occur
• Breakage of glass can occur
• Possible inoculation of broken glass pieces

PARAFILM-ASSISTED MICRODISSECTION TECHNIQUE

After using parafilm the experimenter performed sectioning/slicing (i.e. MICROTOMY) with the help of a very
fine microtome blade
Sharp object cuts’ hazard

After that the cell biologist performed histopathological examination by following a procedure to isolate
Mycobacterium

PBS (Phosphate buffer Saline) solution

Sterile tube
So everything being used is sterilized except the specimen
• Splash hazard
• Aerosols’ generation

Homogenizer (Tissue-grinder) Use

• Splash hazard
• Aerosols’ generation hazard

Petragnani Medium (as a Selective Medium) Preparation

• Micropipette mishandling
• Aerosols’ generation
• Dripping hazard
During colony morphology
• Glass breakage can occur
Then the experimenter performed CULTURING
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 • Breakage of culture tubes
• Spills’ hazard
There are many potential hazards when working with microorganisms
Further Biochemical characterization was done...

This way by identifying the possible risks associated with the procedures used in the article, the instructor
intended to give us an idea of how to perform a procedural risk assessment.

December 9th, 2019 LAB # 6

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ACTIVITY: AUTOCLAVES’ PRESENTATION

The instructor sectioned the class into 5 different groups and assigned a different course topic to each group.
The topic of my group was “AUTOCLAVES”. Our group presented the topic on Monday, 9 th of December 2019.
Later, the group members were asked questions related to AUTOCLAVES’ Biosafety and the instructor marked
our score individually. After completing this course from Canadian Biosafety Program’s website, finally I was
able to receive a certificate of appreciation from THE PUBLIC HEALTH AGENCY OF CANADA, i.e. attached
below

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