Sayad Mahmud Koli

Microbiology Laboratory Layout

Figure: 1
•Documentation room
Purpose:
Documentation room is where all types of documents are recorded in computer or digital format.
It is very important to maintain, compare and finding the issue regarding any product quality.

Apparatus:
Computer
to maintain all data in digital platform.
Printer
To print the document
Router with Internet
For connect with internet
•Media Preparation room
Purposes:
Culture media preparation is one of the routine
tasks common to many microbiology laboratories.
This is true in the food & pharmaceutical industry,
where producers regularly monitor food and
environmental samples for spoilage and
pathogenic microbes as an early indication of
breakdown in processing hygiene.
Common Apparatus:
Petri dish/agar plate
to act as a supporting container to hold the culture
medium in.
Figure: 2
Blood agar plates
(BAP) are made by adding five to ten percent
sheep or horse blood to the nutrient medium.
Nutrient agar grows the largest variety of
microbes, typically fungi and bacteria. It typically
contains nutrients from either beef broth or yeast
extract, depending on what you want to grow.
MacConkey agar plates
(MAC) are a differential type medium made with
bile salts and crystal violet. These additives cause
the agar to only allow growth of Gram-negative
bacteria, while inhibiting the growth of Gram-
positive organisms.
Figure: 3
•Media Preparation room
pH meter Balance
An analytical balance (or lab balance) is a
class of balance designed to measure
small mass in the sub-milligram range.
Use to weight various raw materials for
media preparation

Figure: 4
A pH meter is a scientific instrument that
measures the hydrogen-ion activity in water-
based solutions, indicating its acidity or
alkalinity. The pH meter measures the
difference in electrical potential between a pH
electrode and a reference electrode. It is very
important to maintain the optimum PH of
culture medium.
Figure: 5
•Culture Handling room
Purposes:
separate laminar air flow unit in a separate room for the handling of microbial cultures is a new
trend in pharmaceuticals nowdays. Before sometime all microbiological activities were done on the
same laminar air flow bench including positive cultures, subculturing, sample preparation and
analysis. But now it is changing to have separate area for culture handling and sample analysis.

With this idea in mind, separate culture handling rooms are now used in the pharmaceutical
industries

Common Apparatus :

Laminar Flow cabinet

Laminar air flow cabinets are used in aseptic transfers (also see aseptic techniques ) and aseptic
manufacturing process as well as to facilitate handling of microbiological preparations who’s
sterility is required to be maintained, as well as to prevent cross contamination during material
handling , critical activities which are likely to evolve dust particles like raw material dispensing in
pharma manufacturing process are carried out in a laminar air flow cabinet. (See Fig: 8)
•Incubator room
Incubator
Incubate the prepared culture media at a
specific temperature & times.
Time & temperature
They have a wide variety in incubation
period & Temperature between bacteria
and fungus. In accordance with the
Association for the Advancement of
Medical Instrumentation recommendations
all cultures should be incubated at 37
degrees C for 48 h on suitable culture
media for bacterial growth. For fungal
growth low temperature of 20-25°C is
required and plates are incubated for 72
hours for fungal growth

Figure: 6
•Sterilization room
Purposes:
Sterilization is a critical process in the pharmaceutical industry for the control of microbial
populations microbial populations. While most prevalent in the manufacture of sterile products it
can be used in a variety of settings where microbes have potential impact on patients or products.
The objective of sterilization is to completely destroy or remove all microorganisms (including
spore-forming and non-spore-forming bacteria, viruses, fungi and protozoa) that could
contaminate a healthcare product (for example, those introduced in the manufacturing process
via raw materials, process, equipment, facilities or personnel).
•Sterilization room (Common Apparatus)
Autoclave (Moist heat)
Autoclaves or sterilizers are vessels that use
pressurized steam to eradicate micro-
organisms. Their purpose in the microbiology
laboratory is either to prepare culture media,
reagents and equipment. Or for
decontaminating biohazardous waste
materials.
A general cycle would be anywhere between
3 and 15 minutes, (depending on the
generated heat) at 121 °C (250 °F) at 100 kPa
(15 psi), which is sufficient to provide a
sterility Figure: 7
•Sterilization room (Common Apparatus)
Hot air oven (Dry heat)
Hot air oven is widely used in the
pharmaceutical industry to sterilize the
equipment and other materials that are used in
a laboratory. It is used for delivering the heat
treatment to the product.
The standard setting for a hot air oven is at
least 2 hours at 160 °C (320 °F). A rapid
method heats air to 190 °C (374 °F) for 6
minutes for unwrapped objects and 12
minutes for wrapped objects.
Filtration
Filtration is a great way to quickly sterilize
solutions without heating. Filters, of course,
work by passing the solution through a filter
with a pore diameter that is too small for
microbes to pass through.
Filters can be sintered glass funnels made
from heat-fused glass particles or, more
commonly these days, membrane filters made
from cellulose esters. For the removal of
bacteria, filters with an average pore diameter
of 0.2um are normally used.
Others methods
Ethylene oxide―in a fixed chamber
Liquid chemical sterilants
Radiation (gamma and electron beam)
•Storage room/ Cooling Zone
Purposes: Humidity
Culture media must be stored at the specified Sealed glass and plastic containers are
temperature, under specified conditions and unaffected by normal laboratory humidity.
not longer than the shelf-life periods Opened containers of dehydrated powders
appropriate to each product. The storage will be affected by high humidity. Hot,
conditions and expiry date of each product are steamy media preparation rooms are not
shown on the labels or product inserts but the suitable environments to store containers of
following general rules will help to ensure culture media; particularly containers which
that they are kept in an optimum are frequently opened and closed. An adjacent
environment. When storing products note the cold room or an adequate storage cupboard
shelf life expiry dates on the labels and use are preferable storage areas.
the products in order of their lot/batch
numbers.
Light
All prepared culture media and their
components should be stored away from light
and exposure to direct sunlight should be
avoided at all times.
•Storage room/ Cooling Zone
Temperature and time
The temperature storage conditions of culture
media and their components vary widely. The
following product groupings will help to
differentiate the various requirements.
Culture Media: Sealed, unopened containers
should be stored at room temperature 15-
20°C. Opened containers should have the cap
or lid carefully and securely replaced. It is
important that opened containers are stored in
a dry atmosphere at room temperature. Shelf
life 1 to 5 years.
Prepared Broth Media: Store at 2-8°C. Do
not allow the products to freeze. Shelf life 6
months to 2 years.
Prepared Plates of Culture Media: Poured
plates of agar media are especially vulnerable
to infection, dehydration and chemical
degradation. Aseptic preparation and storage
are essential to protect plates from microbial
infection. Water losses on storage can be
minimised by impermeable wrapping and/or
storage at 2-8°C. Chemical degradation e.g.
oxidation or antimicrobial loss, can be
retarded by protection from light, heat and
dehydration.
It is important, however, to monitor the
storage of prepared plates by quality control
tests so that any deterioration can be detected
and the storage period accurately determined.
Simple weighing tests of fresh and stored
plates will determine the rate of moisture loss.
Weight loss greater than 5% will indicate a
significant loss of water.
•Storage room/ Cooling Zone
Gas Generating Kits:
Store at 2-8°C in a dry place. Do not store
these kits at a higher temperature for long
periods. Shelf life 3 years.

Sterile Reagents:
Store at 2-8°C, except Horse Serum store at -
20 to +8°C.

Susceptibility Discs:
Store at -20°C but keep working stock at 2-
8°C. Shelf life 1 to 2 years.
•Sample testing room
Purposes:
In pharmaceuticals sample testing room used to perform all tests to determine the
quality of a product. Some of the tests performed in pharmaceuticals are discussed
below

Sterility test (To ensure any pharmaceutical product that is fully sterilized. )
Sterilization processes are applied to products in a number of industries, including
food and beverage manufacture, but it is mainly in the pharmaceutical and medical
sectors where the sampling of sterile products for testing remains an important
routine task for microbiologists. Sterility assurance is critical in the manufacture of
many drugs and other medicinal products and is closely regulated worldwide.
•Sample testing room
Sterility testing
(i) fluid mercaptoacetate medium (also known as fluid thioglycolate Medium), which contains
glucose and sodium mercaptoacetate (sodium thioglycollate) and is particularly suitable for the
cultivation of anaerobic organisms (incubation temperature 30–35°C);

(ii) soyabean casein digest medium (also known as tryptone soyabroth), which will support the
growth of both aerobic bacteria (incubation temperature 30–35°C) and fungi (incubation
Temperature 20–25°C).

(iii) Membrane filtration is the technique involves filtration of fluids

Through a sterile membrane filter (pore size 0.45 μm); any microorganism present being retained
on the surface of the filter and the filter is divided aseptically and portions are transferred to suitable
culture media which are then incubated at the appropriate temperature for the required period of
time (7 Days).
•Sample testing room
The Bacterial Endotoxins Test (BET)
is an in vitro assay for detection and quantitation
of bacterial endotoxins, a component of the cell
wall of gram-negative bacteria. The BET is
performed as part of the lot release testing for
medical devices with direct or indirect contact to
the cardiovascular system, lymphatic system, or
cerebrospinal fluid. Injectable pharmaceutical
products must also be tested for bacterial
endotoxins. Routine monitoring of water
systems and incoming materials can help to
ensure that the process does not contribute
endotoxins to the final product.
This assay is also known as the Limulus
Amebocyte Lysate (LAL) test and is sometimes
referred to as the pyrogen test (because bacterial
endotoxins can cause a fever in mammals,
including humans).
Componets:
Amebocyte – Crab blood
Lysate – Separated plasma of Amebocyte
Test procedure:
A BET involves analyzing the liquid sample or
sample extract using Limulus Amebocyte Lysate
(LAL). LAL is a reagent made from the blood of
the crab. In the presence of bacterial endotoxins,
the lysate reacts to form a clot or cause a color
change depending on the technique.
Interpretation:
If clot or precipitation present/colour change –
Endotoxin present in sample
If clot or precipitation absent/colour not changed
– Endotoxin absent in sample
•Sample testing room
•Sample testing room
•Sample testing room
•Sample testing room
•Sample testing room
•Sample testing room
•Sample testing room (Common Apparatus)
•Laminar Flow cabinet •Microscope
•

Figure: 9
•Wire Loop
The loop is used in the cultivation of
microbes on plates by transferring
inoculum for streaking.
•Sprit lamp
•Pipette
•Centrifuge
Separate the contents of a sample
based on their density.
ELISA
Detection of any specific enzyme or
Figure: 8 antibody.