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ACTIVITY 4
ASEPTIC TECHNIQUE
Adapted and edited by Arcibel B. Bautista

LEARNING OUTCOMES:

At the end of the activity, the learner should be able to:

1. Acquaint with the procedures of aseptic transfer of microbiological cultures,;
2. Perform the proper ways of controlling contamination, and

COURSE MATERIAL:

There must be a method of transferring growing organisms from one medium to another
without introducing any unwanted outside contaminants. This method of preventing unwanted
microorganisms from gaining access to samples or cultures is termed aseptic technique.

Any culture of a microorganisms must be regarded as a potential health hazard. The use
of aseptic technique in the transfer of microorganisms from one place to another is vital in
microbiology to ensure:
a. There is no contamination of cultures by microorganisms from the environment;
b. The environment is not contaminated by the microorganisms being handled.

Before starting work, all windows and doors must be closed to prevent draughts. Avoid
sudden movements which might disturb the air. All transfers should take place over a sterile
surface and, particularly when preparing cultures. Ethanol disinfection is recommended because
of its rapid action.

The first requirement of aseptic technique is for all transfer of cultures to be made very
close to a Bunsen burner or alcohol lamp flame. The heat from the flame will cause a draught of
air upwards and so help to prevent microorganisms from the air contaminating an open culture. It
should be noted, however, that if microorganisms are accidentally released through faulty
technique, the up draught will help to disperse them very effectively!

For aseptic technique to be adequate, all items that will come into contact with the
microorganisms must be sterilized properly before and after such exposure. Glassware, media
etc. must be sterilized in an autoclave, pressure cooker or oven as appropriate. When sub-
culturing or inoculating plates, it is often necessary to use equipment such as inoculating loop.

Keeping Sterile Things Sterile

The most common sources of contamination during an experiment are dust, from the
bench top, from the air and from people. This dictates several obvious principles:
1) Keep things covered at all time.
2) Don't touch anything that will come in contact with the culture or any unsterile objects and if
you do touch it sterilize it again before using it.
3) Wipe down the surface around the experiment with alcohol and minimize air turbulence.
4) Avoid talking, singing, whistling, coughing, or sneezing in the direction of things that should
be sterile. Long hair, if not tied back, may be a source of contamination.
5) Maintain a suitable area for preparing, storing, and using sterile media.

GENERAL MICROBIOLOGY laboratory

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Activity 4
ASEPTIC TECHNIQUE

Name: ______________________________________ Score: ________________

Course, Year & Section: _______________________ Date: _________________

In a separate short white bond paper, write all the questions and answers with student
name, course, year and section, and title of the activity. Kindly write the questions given
then answer.

PROCEDURE:

Sterilizing the Inoculating loop and needle

1. Hold the inoculating loop in the area where the wire and the handle meet. Place the
wire at the top of the conical blue flame where heat is at its greatest. Flame the wire
until it glows orange.

2. Slowly move the wire through the flame from handle to tip, allowing each
section to glow orange before moving on.

3. Once the tip of the wire glows orange, remove the loop from the flame.

4. After your loop has been flamed, do not blow on it. Lay it on the countertop, or
touch it to anything other than sterile media or your desired culture unless you’re
done using it; otherwise you have to flame it again.

5. Do the same procedure with the inoculating needle.

QUESTION AND ANSWER:

GENERAL MICROBIOLOGY laboratory

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1. Why is heating the inoculating loop so important in microbiology?

2. Why is it that moist heat is chosen to be the most commonly used in sterilization?

3. What part of the flame is the best part to use to sterilize an inoculating loop or
needle?

Reference:

https://thebiologynotes.com/sterilization-physical-and-chemical-methods/
https://courses.lumenlearning.com/microbiology/chapter/using-physical-methods-to-control-
microorganisms/
https://microbenotes.com/physical-methods-of-sterilization/

GENERAL MICROBIOLOGY laboratory