Laboratoty Biosafety Levels 1

Laboratory Manual in Microbiology and Parasitology
LABORATOTY BIOSAFETY LEVELS
(Submitted by: Rusthia Ann I. Maquirang of BSN 1-2)
Being the lowest of all the biosafety levels, level 1 applies to laboratory settings in which personnel work
with low-risk microbes that pose little to no threat of infection in healthy adults. An example of a microbe that is
typically worked with at a BSL-1 is a nonpathogenic strain of E. coli. This laboratory setting typically consists of
research taking place on benches without the use of special contaminant equipment. A BSL-1 lab also requires
immediate decontamination after spills. Infection materials are also decontaminated prior to disposal, generally
through the use of an autoclave. A BSL-1 lab, is not required to be isolated from surrounding facilities, houses
activities that require only standard microbial practices, such as:
 Mechanical pipetting only (no mouth pipetting allowed)

 Safe sharps handling
 Avoidance of splashes or aerosols
 Daily decontamination of all work surfaces when work is complete
 Hand washing
 Prohibition of food, drink and smoking materials in lab setting, Biohazard signs
Personal protective equipment, such as; eye protection, gloves and a lab coat or gown
Eye Protection Goggles Gloves Lab Gowns

This biosafety level covers laboratories that work with agents associated with human diseases (i.e.
pathogenic or infections organisms) that pose a moderate health hazard. Examples of agents typically worked with in
a BSL-2 include equine encephalitis viruses and HIV, as well as Staphylococcus aureus (staph infections). A BSL-2
laboratory maintains the same standard microbial practices as BSL-1 labs, but also includes enhanced measures
due to the potential risk of the aforementioned microbes. Personnel working in BSL-2 labs are expected to take even
greater care to prevent injuries such as cuts and other breaches of the skin, as well as ingestion and mucous
membrane exposures.
Appropriate personal protective equipment (PPE) must be worn, including lab coats and gloves. Eye protection and
face shields can also be worn, as needed.
Eye Protection Goggles Gloves Lab Gowns Eye Shield

BIOSAFETY LEVEL 3
Again building upon the two prior biosafety levels, a BSL-3 laboratory typically includes work on microbes
that are either indigenous or exotic, and can cause serious or potentially lethal disease through inhalation. An
example of microbes worked with in a BSL-3 includes; yellow fever, West Nile virus, and the bacteria that causes
tuberculosis. The microbes are so serious that the work is often strictly controlled and registered with the appropriate
government agencies. Laboratory personnel are also under medical surveillance and could receive immunizations for
microbes they work with.
Access to a BSL-3 laboratory is restricted and controlled at all times.
Common requirements in a BSL-3 laboratory include:
 All work with microbes must be performed within an appropriate BSC

 Access hands-free sink and eyewash are available near the exit
 Sustained directional airflow to draw air into the laboratory from clean areas towards potentially
contaminated areas (Exhaust air cannot be re-circulated)
 A self-closing set of locking doors with access away from general building corridor.
PPE includes standard personal protective equipment (Gloves, eye protection, and face shield) and respirators might
be required. Also, Solid-front wraparound gowns, scrub suits or coveralls are often required.

Laboratory Respirators Eye Protection Goggles
Eye Shield Coveralls Solid-front wraparound gowns
Scrub suits Gloves

BSL-4 labs are rare. However some do exist in a small number of places in the US and around the world. As
the highest level of biological safety, a BSL-4 lab consists of work with highly dangerous and exotic microbes.
Infections caused by these types of microbes are frequently fatal, and come without treatment or vaccines. Two
examples of such microbes include Ebola and Marburg viruses.
A BSL-4 laboratory is extremely isolated—often located in a separate building or in an isolated and

restricted zone of the building. The laboratory also features a dedicated supply and exhaust air, as well as vacuum
lines and decontamination systems.
Knowing the difference in biosafety lab levels and their corresponding safety requirements is imperative for anyone
working with microbes in a lab setting.
In addition to BSL-3 considerations, BSL-4 laboratories have the following containment requirements:
 Personnel are required to change clothing before entering, shower upon exiting
 Decontamination of all materials before exiting
 A Class III biological safety cabinet
Personnel must wear appropriate personal protective equipment from prior BSL levels, as well as a full body, air-
supplied, positive pressure suit.

Scrub suits Gloves
Full body air-supplied positive pressure suit
REFERENCES

Biosafety Levels 1, 2, 3 and 4. Retrieved February 9, 2020 from https://www.utrgv.edu/ehsrm/programs/lab-

safety/biological-safety-program/biosafety-levels/index.htm.
Full Body Positive Pressure Suit Picture. Retrieved February 9, 2020 from https://www.google.com/search?
q=laboratory+Full+body+positive+pressure+suit&tbm=isch&ved=2ahUKEwjlrO-
Z48PnAhWOzYsBHXKLBJkQ2-
cCegQIABAA&oq=laboratory+Full+body+positive+pressure+suit&gs_l=img.3...3339289.3349851..3350513..
.0.0..0.826.8924.0j9j13j7j1j1j1......0....1..gws-wiz-
img.......0j0i30j0i19j0i5i30j0i8i30.ux4osiW6kbI&ei=pZ0_XqXGE46br7wP8paSyAk&bih=608&biw=1366#imgr
c=Jhs9GyeHREt5zM
Laboratory Coveralls Picture. Retrieved February 9, 2020 from https://www.google.com/search?

q=laboratory+Coveralls&tbm=isch&ved=2ahUKEwiF8KKV48PnAhVF3pQKHXqkB14Q2-
cCegQIABAA&oq=laboratory+Coveralls&gs_l=img.3...0.0..2258...0.0..0.0.0.......0......gws-wiz-
img.1pAQbdv3l6Y&ei=m50_XsW2KcW80wT6yJ7wBQ&bih=608&biw=1366#imgrc=fobOaEQeJhTT5M
Laboratory Scrub Suits Picture. Retrieved February 9, 2020 from https://www.google.com/search?

q=laboratory+Scrub+suits+&tbm=isch&ved=2ahUKEwi9odOy4MPnAhVGw4sBHW1gBVAQ2-
cCegQIABAA&oq=laboratory+Scrub+suits+&gs_l=img.3...677474.690717..691030...0.0..0.1130.10257.2-
8j3j3j4j3j1......0....1..gws-wiz-
img.......0i67j0j0i30j0i19.2dATTGXuTTY&ei=s5o_Xv37O8aGr7wP7cCVgAU&bih=608&biw=1366#imgrc=Q3
Dfhowq00fCmM
Biosafety Levels Picture. Retrieved February 9, 2020 from https://www.google.com/search?

q=Biosafety+level+1&sxsrf=ACYBGNTnfZ-
T1lu8UgvthDjsESU1r0OJ_A:1581220374152&source=lnms&tbm=isch&sa=X&ved=2ahUKEwj92Nn1yMPnA
hWb-2EKHSYBCDEQ_AUoAXoECAwQAw&biw=1366&bih=657
Solid-front wraparound Laboratory Gowns Picture. Retrieved February 9, 2020 https://www.google.com/search?

q=Solid-
front+wraparound+laboratory+gowns&tbm=isch&ved=2ahUKEwjNlbij4MPnAhUIWpQKHdDhBDgQ2-
cCegQIABAA&oq=Solid-
front+wraparound+laboratory+gowns&gs_l=img.3...24592.27693..28193...0.0..0.0.0.......0....1..gws-wiz-
img.nboiha3t4pc&ei=lJo_Xs37BIi00QTQw5PAAw&bih=608&biw=1366#imgrc=6Iabkl2x_h_RuM
Lab Respirator Picture. Retrieved February 9, 2020 https://www.google.com/search?

q=Lab+respirators&sxsrf=ACYBGNSLvIJs38q-DLK-
S8qKcjHydxMKPg:1581225909302&source=lnms&tbm=isch&sa=X&ved=2ahUKEwiztojF3cPnAhWBMd4KH
QWHDTMQ_AUoAXoECAwQAw&biw=1366&bih=608#imgrc=Vd4Pc1PVc1l7IM
EQUIPMENTS, APPARATUS, AND TOOLS USED IN MICROBIOLOGY AND PARASITOLOGY
(Submitted by: Rusthia Ann I. Maquirang of BSN 1-2)

Microscope Slide
A microscope slide is a long thin piece of glass that specimens are placed on for study under a microscope.
It is easier to manipulate the slide, rather than the specimens, into the best position for viewing, as many are fragile
and microscopic. Slides normally measure three inches by one inch and can be made of clear plastic in addition to
glass. Some slides have small depressions to hold liquid for wet mounts. A wet mounted slide is one where liquid,
usually a stain, is placed over the specimen. Stains help you see the organism better by highlighting the individual
features.
Test tube
Also known as a culture tube or sample tube, is a common piece of laboratory glassware consisting of a
finger-like length of glass or clear plastic tubing, open at the top and closed at the bottom. Test tubes are usually
placed in special-purpose racks.
Petri Dish
A Petri dish is a shallow nesting dish made of glass or plastic that is typically used in laboratories. These
dishes also have clear lids that allow scientists to view the contents without removing the lid. Scientists use them to
grow bacteria, mold and other microscopic organisms that they can then examine under a microscope.
Inoculation loop

An inoculation loop, also called a smear loop, inoculation wand or microstreaker, is a simple tool used
mainly by microbiologists to pick up and transfer a small sample (inoculum) from a culture of microorganisms, e.g.
for streaking on a culture plate.
Pipettes
Also called pipets or chemical droppers, are small tubes of glass or plastic used to transfer a
measurable amount of liquid from one container to another. They come in two forms: volumetric pipettes, used to
transfer a single specific volume of liquid, and measuring pipettes, used to transfer varying, measured volumes.
Pipettes in their current form appeared in the 1970s, to replace the old and dangerous practice of mouth
pipetting, where scientists would transfer liquids in the lab using straws and suction from their own mouths,
regardless of potential hazards.
Hot Air Oven for Sterilization

It is used for sterilization of glassware’s, such as test tubes, pipettes and petri dishes. Such dry sterilization
is done only for glassware’s. Liquid substances, such as prepared media and saline solutions cannot be sterilized in
oven, as they lose water due to evaporation. The glassware’s are sterilized at 180°C for 3 hours. An oven has a
thermostat-control, using which the required constant temperature can be obtained by trial and error. The thermostat
dial reading is approximate and the exact temperature is read by introducing a thermometer into the oven or on a
built-in L-shaped thermometer.
In a modern oven, there is a digital temperature display and automatic temperature controller to set the
desired temperature easily. Time is set by a digital timer. After loading the glassware’s, the door is closed and oven
switched on. The required temperature is set. After the oven attains the set temperature, the required time of
sterilization is set on the timer. The oven switches off automatically after the set time. The oven is opened, only after
its temperature comes down near to room temperature. Otherwise, if the door is opened, while the inside of the oven
is still very hot, cold air may rush in and crack the glassware’s.
Drying Oven
For preparation of certain reagents, the glassware’s, after proper cleaning and rinsing with distilled water,
are required to be dried. They are dried inside the drying oven at 100°C till the glassware’s dry up completely.
Autoclave
Autoclave is the nucleus of a microbiology laboratory. It is used not only to sterilize liquid substances such
as prepared media and saline (diluents) solutions, but also to sterilize glassware’s, when required. It has the same
working principle as a domestic pressure cooker. The maximum temperature that can be obtained by boiling water in
an open container is 100°C (boiling point of water). This temperature is sufficient to kill only the non-spore formers,
but it is difficult to kill the spore-forming bacteria at this temperature, as they escape by forming heat resistant spores.
It takes very long time to kill the spores at this temperature. On the other hand, when water is boiled in a closed
container, due to increased pressure inside it, the boiling point elevates and steam temperature much beyond 100°C
can be obtained. This high temperature is required to kill all the bacteria including the heat resistant spore-formers.
Steam temperature increases with increase in steam pressure.

Microbiological Incubator
Profuse growth of microbes is obtained in the laboratory by growing them at suitable temperatures. This is
done by inoculating the desired microbe into a suitable culture medium and then incubating it at the temperature
optimum for its growth. Incubation is done in an incubator (Figure 3.7), which maintains a constant temperature
specifically suitable for the growth of a specific microbe. As most of the microbes pathogenic to man grow profusely
at body temperature of normal human being (i.e. 37°C), the usual temperature of incubation is 37°C.
Fridge (Refrigerator)
It serves as a repository for thermo labile chemicals, solutions, antibiotics, serums and biochemical reagents
at cooler temperatures and even at sub-zero temperatures (at less than 0°C). Stock cultures of bacteria are also
stored in it between sub-culturing periods. It is also used for the storage of sterilized media, so as to prevent their
dehydration.
Electronic Top-pan Balance

It is used for weighing large quantities of media and other chemicals, where precise weighing is not of much
importance.
Electronic Analytical Balance

It is used to weigh small quantities of chemicals and samples precisely and quickly.

Distilled Water Plant

Water is used in the preparation of media and reagents. If the media are prepared using tap water, the
chemical impurities present in it may interfere with the growth of the microorganisms in the media. Moreover, the
higher is the bacteria content of the media, the longer is the time required for their sterilization and greater is the
chance of survival of some bacteria. Distilled water, though not bacteria- free, contains less number of bacteria. That
is why; it is preferred in the preparation of microbiological media. It is also used in the preparation of reagents,
because the chemical impurities present in tap water may interfere with the proper functioning of the reagent
chemicals. As manufacture of distilled water by Liebig condenser is a time-taking process, in most laboratories, it is
prepared by ‘distilled water plants’. Usually a distilled water plant is made of steel or brass. It is also called distilled
water still. It has one inlet to be connected to the water tap and two outlets, one for distilled water to drop into a
container and the other for the flow out of hot cooling water into the sink. The still is installed on the wall. It is heated
by in-built electric heating elements (immersion heater).
Microscope
Microscope consists of two lens called eye lens and objective lens, objective lens is lens kept behind object
and eye lens is keep on the top of microscope i.e. on the place through which we look, firstly the object is placed
behind the objective of microscope which is turned into virtual, erect and magnified image, later this image is thought
to be the object for the eye lens and this objects forms real, inverted and magnified image.

Laminar Flow Hoods

Laminar flow hoods are essential machines to medical research. They are an enclosed worktop that
scientists can keep entirely sterile. In the hoods, scientists can do various treatments on cells or experimental
animals, keeping both the test subjects safe from outside contaminants, and scientists safe from possible threats
inside the hood, like virus particles. However, they require strict maintenance to ensure the region inside the hood
remains sterile and safe.
REFERENCES
UC Irvine. (2020).Volumetric Flasks and Pipets. Retrieved February 10, 2020 from https://sciencing.com/differences-
lab-glassware-8091302.html
Samiksha S. (2020). http://www.yourarticlelibrary.com/micro-biology/29-equipments-every-microbiology-laboratory-

should-have/2647
Ask Media Group, LLC. ( 2020). Petri dish. Retrieved February 10, 2020 from
https://www.reference.com/science/function-petri-dish-ba4df5d1a96076e3
Microscope Slide. Retrieved February 10, 2020 from https://sciencing.com/functions-glass-slide-cover-slips-

6680506.html
Test tube. Retrieved February 10, 2020 from https://en.wikipedia.org/wiki/Test_tube
Thompson BF, Thompson R (2012). Illustrated Guide to Home Biology Experiments. February 10, 2020 from
https://en.wikipedia.org/wiki/Inoculation_loop
Deepshikha. (2019). 8 Basic Laboratory Equipment Used in Microbiology | Biology. Retrieved February 10,2020 from
http://www.biologydiscussion.com/micro-biology/laboratory-equipment/8-basic-laboratory-equipment-used-
in-microbiology-biology/8575


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Laboratory Manual in Microbiology and Parasitology