Beruflich Dokumente
Kultur Dokumente
By
NIK ABDUL HAFIIDZ BIN NIK ABDUL MALEK
2014688106
SAIFUL ANWAR BIN NAZURALLAH
2014626794
AHMAD AHKMAL BIN MASRUDIN
2014631198
MOHAMMAD ARSHAD BIN RASHID
2014683386
SEPTEMBER 2015
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TABLE OF CONTENTS
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Chapter 1
Hazard Identification
In Malaysia, laboratories can be found in universities, companys research and development
and government institute. The laboratory environment can be a hazardous place to work.
Laboratory workers such as students are exposed to numerous potential hazards including
chemical, biological, physical and radioactive hazards, as well as musculoskeletal stresses.
Laboratory safety is governed by numerous local, state and federal regulations. The reduction
of hazards improves the safety and quality of life for human and for the environment, these
hazards can be minimized through researches and improve safety procedure (Maaza, 2008).
Therefore performance of risk assessment of hazardous chemical and biohazard is an
important process to organize the management of hazards of these chemicals and biohazard.
This study was conducted to assess the risk from genetic engineering laboratory course in
chemical and bioprocess engineering program.
1.1
Hazard is defined anything for example condition, behaviour, situation, practice that has the
potential to cause harm, including disease, death, injury, environmental, property and
equipment damage. A hazard can be a thing or a situation. (Western Sydney University, 2015)
Biological hazards, also known as biohazards, are organic substances that pose a
threat to the health of humans and other living organisms. Generally speaking, biological
hazards include pathogenic micro-organisms, viruses, toxins (from biological sources),
spores, fungi and bio-active substances. Biological hazards can also be considered to include
biological vectors or transmitters of disease. Outside the health arena, biological hazards
include substances that cause social and economic disruption, property damage and
environmental degradation, such as insect plagues or infestations. Worldwide, it is estimated
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that around 320 000 workers die each year from communicable diseases caused by workrelated exposures to biological hazards (Safe Work Australia, 2011).
1.2
Identify Hazards
Identify hazards is the first step in risk management step in the workplace. A hazard is anything
including work practices or work practice or procedures that have the potential to harm the
health and safety of a person. The student is responsible for identifying hazards which may
be present in the workplace. Laboratory safety requires knowledge of laboratory procedures,
equipment, and reagents, as well as constant watchfulness for danger. One careless student
can cause accident to other student. Hazard is classified as below:
Electrical hazard are sources of danger from electrical equipment that can cause
electrical shock Example of Electrical equipment placement of cords, wet areas, hot plates.
Potential impact electrocution and fire;
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1.3
blood
body fluids
urine
bacterial smears
bacterial cultures
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1.3.1
In this experiment there is no biohazard involve. However, the swab use to obtain human
cheek cell should be clean and avoid any bacteria and other hazardous material enter the
mouth. The chemical use in this experiment cool isopropanol and soap which is not hazardous
to heath but are highly Flammability. Electrical equipment use is centrifuge which should be
handle carefully. Physical hazard may occur according to statement above. the aim of this
experiment is to isolate gene.
The experiment involve the use of microbacteria E. Coli which is classify under biosafety level
1 and risk group 1. E. Coli is a microorganism that is unlikely to cause human or animal disease
but the high concentration of microorganism that enter the body may cause diarrhea. In this
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experiment, the E.coli cell was transform by adding new foreign gene which is pGlo to observe
its bacterial transformation. The newly transformed bacteria should be keep safe and avoid in
contact with human body. The aim of experiment is to develop an understanding of the
biological process of bacterial transformation by the pGLO plasmid DNA. Physical and
chemical hazard may occur according to statement above.
Figure 3: transformed bacteria (E. Coli) is covered to avoid contaiminantion to sample and
avoid exposure.
1.3.3
Dna
The aim of this experiment is to separate and sort a large group of DNA molecules by size,
determine the size of each molecule separated and develop an understanding of the role of
restriction enzymes and agarose gel electrophoresis to cut and size DNA. The possible
biohazard in this experiment is the enzyme use in this experiment. There are probability that
when enzyme enters into human body it may cause sickness. The major hazard in this
experiment is during gel electrophoresis, the buffer used which contain electric current should
be keep safe and do not touch the buffer liquid . it may cause electrical shock. Physical hazard
may occur according to statement above.
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Figure4: Agarose gel before (left) and after (right) electrophoresis procedure
1.3.4
In this experiment, the procedure quite familiar to the restriction endonuclease digestion and
gel electrophoresis of dna lab only the sample where change for the electrophoresis. The
major hazard involve in this experiment is during electrophoresis. The dna sample of the
experiment is consider non-biohazard. No microbial use in this experiment. Physical, chemical
and electrical hazard should be avoided.
Figure 5: The PCR tube containing DNA sample, mastermix and primers.
1.3.5
The aim of this experiment is study the basic ELISA procedure to measure the concentration
of antibodies or antigens in solutions. In this experiment, the potential biohazard is sample
taken from student which is human saliva. It may contain some bacteria. When exposed to
another student it may cause low risk of infection depends on that type of virus or infection
that student carries.
(A)
(B)
Figure 6: Result of ELISAs test shows patients with antigen (left) and patients without antigen
(right).
--------------------------------------------------------------------------------------------------------------------------Chapter 2
The Biological Risk Assessment
Our laboratory manager and laboratory assistant use risk assessment to alert their
staffs to the hazards of working with infectious agents and to the need for developing
proficiency in the use of selected safe practices and containment equipment. This procedure
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used in our UiTM Bioprocess Laboratory. Successful control of hazards in the laboratory also
protects persons not directly associated with the laboratory, such as other occupants of the
same building, and the public. This follow the Occupational Safety & Health Act 1994,
USECHH 2000 Regulation.
The Lecturer or laboratory assistant must consult with a Laboratory manager to ensure
that the laboratory is in compliance with established guidelines and regulations. When
performing a risk assessment, it is advisable to take a conservative approach if there is
incomplete information available. Factors to consider when evaluating risk include the
following:
Pathogenicity
Route of transmission
Agent stability
Infectious dose
Concentration
Origin
Medical surveillance
In this assignment I need to categorize the risk group, biosafety level and safety containment
level based on the experiment that we have done in genetic laboratory experiment. We need
to ensure that all the experiment follow the guideline.
1) RISK GROUPS
In many countries, including the United States, biological agents are categorized in Risk
Groups (RG) based on their relative risk. Depending on the country or organization, this
classification system might take the following factors into consideration:
Other factors
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It is important to understand that biological agents are classified in a graded fashion such that
the level of hazard associated with RG1 being the lowest and RG4 being the highest. EHS
Biosafety follows the NIH Guidelines categorization of Risk Groups as follows:
RG1 Are not associated with disease in healthy adult humans or animals
RG2 Are associated with disease which is rarely serious and for which preventative
or therapeutics is often available
RG3 Are associated with serious or lethal human disease for which preventative or
therapeutics may be available
RG4 Are associated with lethal human disease for which preventative or therapeutics
are not readily available
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2) BIOSAFETY LEVELS
In contrast to Risk Groups, Biosafety Levels (BSL) prescribe procedures and levels of
containment for the particular microorganism or material (including Research Involving
Recombinant or Synthetic Nucleic Acid Molecules). Similar to Risk Groups, BSL are graded
from 1 4. Detailed descriptions of containment practices and biosafety levels can be found
in our laboratory guidelines.
The majority of experiments done at UiTM Bioprocess Laboratory involves Biosafety Level
1 and 2 (BSL-1) & (BSL-2) practices. BSL-2 containment and practice is suitable for work with
agents that are infectious to humans or animals where exposure may result in limited to
moderate disease. The routes of exposure to these agents are typically through cuts and
breaks in the skin, ingestion, and splashes to the mucous membranes (eyes, nose, and
mouth). These agents or materials include:
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in this experiment, we can conclude that it can be put under risk group 1, containment level 1
and biosafety level 1.
EXPERIMENT 2 BACTERIAL TRANSFORMATION
This is the abstract of the experiment. Transformation is basically the process in which a
bacterium takes up an exogenous DNA and expresses its genes, resulting in changes to its
traits. One of the commonly used genes to demonstrate bacterial transformation is a gene that
codes for Green Flourescent Protein (GFP). When exposed to long ultraviolet (UV) wave light,
it emits bright green light that is visible in bacteria transformed by plasmid that contain the
genes encoding GFP. In this experiment, bacteria were transformed using transformation
solutions and heat shock procedures. The bacteria used in this experiment is a commonly
used bacteria used for genetic transformation is the Escherichia coli, or E. coli. And the
plasmid used to transform the bacteria is the pGLO plasmid, which encodes a gene for GFP
and a gene for resistance to the antibiotic ampicillin. These characteristics of the plasmid
provide a viable screening and selection process for transformed bacteria. Adhering to the
objectives of the experiment, an understanding of the biological process of bacterial
transformation by the pGLO plasmid DNA was developed and the acquired phenotypic trait of
GFP exhibited by transformed bacterial was observed.
In this experiment, we deal with Escherichia coli, or E. coli which is the microorganism
that classified under risk group 2. In order to deal with the bacteria, we must follow the
containment level 2, thus this experiment also can be categorize under biosafety level 2
EXPERIMENT
RESTRICTION
ENDONUCLEASE
DIGESTION
AND
GEL
ELECTROPHORESIS OF DNA
This is the abstract of the experiment. Restriction enzymes are like scissors for DNA. They
are biomolecules that restrict (cut), DNA at specific sites. Agarose gel electrophoresis is a
powerful separation method frequently used to analyse DNA fragments generated by
restriction enzymes. Electrophoresis refers to the migration of a charged molecule through a
restrictive matrix, or gel, drawn by an electrical force. This experiment aimed to separate and
sort a large group of DNA molecules by size and determine it, as well as to develop an
understanding of the role of restriction enzymes and agarose gel electrophoresis to cut and
size DNA. Agarose gel electrophoresis was successfully used to separate and sort a large
group of DNA molecules by size. This is apparent through the different bands showed by the
agarose gel after the electrophoresis. The size of each molecule separated was determined
using two methods: direct gel examination and standard curve of the semi-log graph.
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In this experiment, we use restriction enzyme as the main microorganism. The restriction
enzyme not harmful if only at our skin but it may be harmful if it is inserted inside the human
body. This experiment also deal with high voltage and high temperature equipment. From what
we have done in this experiment, it can be categorize under risk group 2 that lead to
containment level 2 and also can be put under biosafety level 2.
EXPERIMENT 4 THE ISOLATION OF GENETIC MATERIAL BY PCR
This is the abstract of the experiment. This experiment aims to demonstrate and understand
the techniques of PCR, analyze and evaluate its results and also to determine its sensitivity.
This was done in the need of DNA fingerprinting or DNA profiling for a crime scene
investigation. The sample from the crime scene as well as the DNA for a group of suspects
was put under the procedures of PCR. Following the procedure, the amplifying process was
easily achieved.
In this experiment, there are no biological harmful effect because all the sample was already
prepared by lab assistant. The risk that may involve is when we want to transfer the genetic
from a tube to another and also we need to deal with high temperature and high voltage
equipment. Therefore we can categorize them under risk group 2 that lead to containment
level 2 and also can be put under biosafety level 2.
EXPERIMENT 5 ELISA
This is the abstract of the experiment. The Enzyme linked immunosorbent assay (ELISA)
uses antibody to detect the presence of a disease agent such as viruses, bacteria and
parasites in the blood or other body fluid. This experiment demonstrates the application of
ELISA in real situation. The enzyme, HRP is essential to present the positive result of the
assay where it oxidizes the colorless substrates into a blue solution. The blue color indicates
that there are antigens present in the serum and the antibodies have successfully bound onto
them. This experiment was fairly successful since all the wells gave out different result for
different samples, but accordingly to the theory. Moreover, the results that was taken into
consideration was only the color changes that occurred within 5 minutes. Therefore, the results
are deemed acceptable.
In this experiment, the sample used is unknown but it is under risk group 2 microorganism.
These experiment also can detect the harmful microorganism such as virus SARS or even
HIV. That virus can be categorize under risk group 4. In our level although the sample are
unknown or maybe in form of GMO, the risk group involve is in group 2. In order to handle it,
we must follow the biosafety level 2 or 3 and the containment level 2.
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Chapter 3
Biohazard Equipment
While handling experiment in the laboratory, many safety precaution needs to follow by person
to make sure each experiment finish with success. Before uses every equipment in the
laboratory, each person needs to read the sign on that particular equipment. These show how
to handling without any problem happens.
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Every laboratory also need smoke alarm to detect fire before huge fire happen and
damage chemical. Explosion in laboratory will damage the entire chemical in the storage and
this will increase exposure of poison smoke into person. Exposures of chemical are very
dangerous because it will widely expose through the air and move about 2km and more around
the accident place.
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When any accidents happen in the laboratory, every person must take immediately
precautions to help themselves. Emergency maps must shown at every building to make sure
each person move out from that building in safety condition to the assembly point. Follow the
exit sign that shown on the laboratory door to the assembly point.
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NAME OF
EXPERIMENT
CHEMICAL USES/
MICROORGANISM
USES
Elisa
Phosphate
buffer
Genes in the
Lysis buffer
bottle
Isopropanol
The isolation of
Orange G
genetic
loading dye
material by
TAE buffer
PCR
Bacterial
Escherichia
transformation
coli
Calcium
Chloride
Restriction
Buffer
endonuclease
solution
digestion and
EcoRI
gel
Pstl
electrophoresis
HindIII
of DNA
3.1
LAMINAR
FLOW
HOOD
MASK
LAB
COAT
GOGGLES
GLOVES
ELISA uses antibody to detect the presence of a disease agent such as viruses, bacteria and
parasites in the blood or other body fluid. ELISA have been used in pregnancy test, disease
detection in people, plant and animals, detecting illegal drug use, test for indoor air quality and
test to determine whether the food is labelled accurately. In general, an antigen is any
molecule that can provoke the immune system to respond. This immune response is specific
to a type of antigen (Panigrahi, B. K., Abraham, A., & Das, S., 2010). There are two options of
ELISA detection which are direct detection and indirect detection.
Goggles
Lab Coat
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Gloves
Masks
Figure 18: Safety Equipment use in lab
3.2
Protein synthesis involves two main steps: transcription and translation. The DNA plays an
important role as the first molecule that is needed in the process. The letters of the nucleotides
encoded in the DNA is copied by enzymes, basically forming 3 types of RNA; the messenger
RNA (mRNA) the transfer RNA (tRNA) and the ribosomal RNA (rRNA) (Brown, 2010). These
experiment shows DNA extraction and precipitation and to observe a collection of DNA
strands. DNA that been collecting from human body was check cells to use in these
experiment. To break open the cells to release the DNA, lysis buffer contains detergent, will
been used. Then, to make the DNA visible cold alcohol is used. Using isopropanol, the DNA
is able to get out of the solution in which it is in, and form clumps to differentiate itself. To
handle both of these chemicals, mask, goggle, gloves and lab coat must been used while
handling during the experiment.
3.3
It is a polymerase chain reaction (PCR) technique that determines the alleles present at
different short tandem repeats (STR) loci within a genome in order to use DNA information to
identify individuals (Andreas, Nicole and Dimitri, 2004). From this experiments, the objective
to demonstrate and understand the techniques of PCR, to evaluate the results of PCR and to
determine the sensitivity of PCR.
3.4
Bacterial transformation
Genetic transformation has given rise to many successful findings in terms of medicinal,
agricultural and bioremediation aspects. For example, agriculture sector have been modified
to have better resistance towards pests and other damaging factors, as well as to improve its
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growth. Bacteria have also been transformed to assist in bioremediation processes, in terms
of pollutant absorption and elimination. E.coli will be used for genetic transformation. Like all
bacteria, E. coli does not have any nuclear membrane or a nucleus, thus indicating that all of
the genes required for its survival are only found in a single chromosome (Karp and Pruitt,
1996). To handle E.coli, mask, gloves, goggles, lab coat and laminar flow hood was been used
to prevent E.coli from death and affect human body from get diarrhea.
Figure 19: Laminar Flow Hood used to avoid exposure to the microorganism
3.5
Restriction enzymes are like scissors for DNA. They are biomolecules that restrict (cut) DNA
parts. Agarose gel electrophoresis is a powerful separation method frequently used to analyze
DNA fragments generated by restriction enzymes (Mozayani, 2011). This experiment aimed
to separate and sort a large group of DNA molecules by size and determine it, as well as to
develop an understanding of the role of restriction enzymes and agarose gel electrophoresis
to cut and size DNA. All the restriction enzyme uses was transferred into tubes. Then, DNA
that through process of cutting fragment by restriction enzyme will move to electrophoresis
process.
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Chapter 4
Emergency Response Procedure/ Report/ Accident
From our interview with the secretary of safety and health committee of FKK, Miss Hajatun
Rabani binti Ahmad Razif. Till today, there are only one major accident which are the
laboratory ceiling dropped but it did not cause any injuries. Most common accident occurs in
the laboratory are broken beakers which mostly caused by mishandling. There are no serious
injury due to accidents in the laboratory that are reported until today.
Provided below is the form used to report any accident occurs in the laboratory, the
first person that need to be informed first are the lab assistant to measure the severity of the
accident for further action needs to be taken. From the form, we can see that any accident are
treated very seriously in spite of their severity. This is to comply with the FKK safety and policy
which is to continually improved compliance with OSH legislations and effective
implementation of FKK UiTM OSHMS by achieving and maintaining certification to OHSAS
18001:2007 and MS1722:2011.
Also provided below is the standard emergency procedure when fire occurs in the
laboratory, this procedure must be shown and followed by all personnel that works in the
laboratory to prevent chaos and reduce risk of injury in case of fire occurs.
Chapter 5
Recommendation For Improvement In Fkk Laboratory
Up till today, there are no accident that have caused serious injuries or illness in FKK
laboratories. This proves that the preventive and proactive action taken by the management
is successful. However, there is always room for improvement. There are few suggestion that
can be taken into consideration by the management in order to increase the safety of the
laboratory.
Place a printed policy of statement of environment, health and safety from the faculty
Organize a talk on safety which involves management, employees, faculty, staff and
students.
Make it a requirement for new employees and students to go through safety orientation
to expose them to the equipment they will use and explain the hazard of the chemical
that will be use in the future.
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Ensure students to read safety manual of the equipment that they will be using along
with the lab manual.
Conduct announced lab inspection to ensure all equipment are in the best condition at
all time.
Students must not be allowed to work alone in the lab without approval from the staff.
Every experiment must be monitored until finish unless they have failsafe.
Lecturer must explain the hazard of the chemical that will be used and PPE that are
required before conducting experiments.
References
A & A SCIENTIFIC RESOURCES SDN BHD, 2011. Malaysias 1st University Affiliated
Environmental
Laboratory.
Retrieved
on
27
September
2015
from
http://www.environment.com.my/about/
Andreas Manz, Nicole Pamme and Dimitri Lossifidis (2004), Bioanalytical Chemistry; Imperial
College Press,
Anonymous 2014. 40 steps to improve lab safety. Retrieved September 27, 2015, from
http://safety.nmsu.edu/programs/lab_safety/steps_to_lab_saf.htm
Brown, T. (2010). Gene cloning and DNA analysis: an introduction: Wiley-Blackwell.
FKK, 2012. Ohsasform/Accident Incident Investigation Report retrieved on September 27,
2015, from
http://fkk.uitm.edu.my/v1/Ohsasform/Accident
Incident Investigation
Report (1).pdf
Karp, G., & Pruitt, N. L. (1996). Cell and molecular biology: concepts and experiments: John
Wiley & Sons New York.
Maaza Yousuf Abdulah Mohamed, 2008. Risk Assessment Of Hazardous Chemicals A Case
Study In Chemistry Research Laboratories Of USM. University Sains Malaysia
Mozayani, A. (2011). The forensic laboratory handbook procedures and practice: Springer
Science+ Business Media.
Safe Work Australia, 2011. National Hazard Exposure Worker Surveillance Exposure to
biological hazards and the provision of controls against biological hazards in Australian
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workplaces
retrieved
on
26
september
2015
from
http://www.safeworkaustralia.gov.au/sites/SWA/about/Publications/Documents/571/N
HEWS_BiologicalMaterials.pdf
Western Sydney University, 2015. Hazard Identification, Risk Assessment and Control
Procedure.
Retrieved
on
26
September
2015
from
http://www.uws.edu.au/__data/assets/pdf_file/0020/12917/12917_Hazard_Identificati
on,_Risk_Assessment_and_control_Procedure.pdf
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Appendix
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