Lesson 11: ASEPTIC TECHNIQUE Flashcards
The main objective of aseptic technique
to eliminate contaminants from
work surfaces, solutions and other sources in the laboratory.
Elements of aseptic environment
- Quiet area
- maintained work surface
- Personal hygiene
- Sterile within packages Reagents and media
- Cautious handling of cell cultures
Contamination can be avoided by
- regular checking of culture
- avoidance of use of antibiotics for at least a part of incubation period to reveal cryptic contaminations
- always use sterile reagents and materials
- exclusive use of media, materials for different cell lines
- strict observation of standard sterile technique at all times.
in the absence of a laminar flow or sterile room, a quiet area with little or no traffic and no other activity should be picked. The area should be kept clean and not contain equipment other than that related to culture. Non-sterile activities should not be
Quiet Area
maintain clean and tidy work surface by
a. completely clear surface
b. swab surface with 70% alcohol before, between and after procedures
c. place onto the work surface materials necessary for a certain procedure
d. arrange work area systemically for easy access of items used and wide clear space to work on
e. work within range of vision
f. mop up any spillage immediately and swab area with 70% alcohol, and
g. when finished, remove all materials from the work surface and swab with 70% alcohol.
hand washing is encouraged to reduce bacterial count/adherent microorganisms on the skin that would otherwise likely to blow onto your culture and cause contamination. Surgical gloves may be worn and swabbed frequently, and disposed properly after procedure. In addition, long hair are tied back or kept in a cap, avoid talking when doing a procedure if not working in a laminar flow hood, or avoid doing any culture procedures when having any respiratory symptoms.
Personal Hygiene
commercial reagents and media are ensured to be sterile within packages. So it is advisable to always swab its external surface with 70% alcohol before use if possible.
Reagents and media
Be cautious when handling cell cultures from other laboratories as they may be contaminated from the source or during transit. Newly arrived cell lines should be handled separately from the rest of the stocks and avoid antibiotics until they are shown to be uncontaminated.
Cultures
Do this in the surface with 70% alcohol before, between and after procedures. Do this to other materials if applicable before use.
Swabbing
Sterile Handling
Swabbing
Capping
Flamming
Handling bottles and flasks
Pipetting
Pouring
screw caps should be washed thoroughly and covered with aluminum foil
to protect neck of the bottle from dust.
Capping
Doing this on the neck of bottles and screw caps or other materials before and
after opening, before and after closing it will avoid contamination.
Flaming
when working on an open bench, bottles and culture flasks should not be kept vertical when opened and kept at an angle as shallow as possible during manipulations to avoid contamination
Handling Bottles and Flasks
should be avoided as much as possible to prevent generation of a bridge of liquid between the outside of the bottle and the inside, which may permit contamination to enter the bottle.
Pouring
standard glass or disposable plastic is the easiest way to manipulate liquids. Doing this with your mouth should be avoided to avoid mycoplasmal contamination and prevent hazards to the operator.
Pipetting
