Rules & procedures for Tissue
Culture Lab

Only authorised users may use the TC
facilities

Only tissue culture can be performed in the
TC lab. There are to be absolutely NO animal
dissections or other lab work carried out in the
TC lab

Use your own reagents – do not use others’
reagents unless you ASK THEM FIRST!

Clean up any mess you make! The tissue
culture room runs smoothly based on an honesty
system. To leave the TC room in a mess is selfish
and inconsiderate of other users.

Users who do not comply with the rules will
be banned in the future from using TC facilities.
Use of the Class II hood
1) All usage must be logged in the logbook
2) Open the front panel and turn on the air flow &
light. Let the air flow stabilise
3) Use 70% ethanol to sterilise the hood prior to
use, as well as any other equipment to be used
inside the hood (ie: tip boxes, pipettes)
4) Dispose of all tips/glass pipettes, liquid waste,
contaminated/non-contaminated waste
appropriately (see waste disposal below)
5) After use, wipe the inside of the hood and used
equipment with 70% ethanol. Clean up all spills
6) Turn off the hood and wait for the air flow to
fully stop before closing the front panel
7) Suction 5-10mL of bleach & empty liquid waste
container into sink with copious amounts of
water. Add 10mL bleach to the bottle before
putting it back into its place
Waste disposal
Solid waste
Glass pipettes,
sharps, pipette
tips
Contaminated dry
waste/tissue culture
plates/dishes
Non-contaminated
dry waste
Sharps
container
*No liquids
Autoclave waste bin
Normal bin
Liquid waste
TC media
Bleach &
discard into
sink with lots
of water
Solvents
Halogenated waste
container (L9-14)
Non-halogenated
waste container (L9-14)
Eg: halothane, methylene chloride,
chloroform, carbon tetrachloride,
trichloroethane, trichloroethylene
Eg: acetone, methanol, ethanol,
toluene, xylene, acetonitrile, benzene
etc
Miscellaneous waste
Trizol
Ethidium bromide
Trizol waste
container (L9-14)
Ethidium bromide waste
container (L9-15)
Different types of heavy metal solutions, mineral acids, inorganic bases,
oxidizers & reactive wastes should be stored SEPARATELY & should never
be mixed with anything else! DO NOT pour any solvents and toxic
chemicals down the sink! If you are unsure about disposal of
anything, please ask Helen (or Tony if Helen is absent).
Use of the CO2 incubator






Sterilise your hands with 70% ethanol prior to taking out
your cultures
Before putting your cultures inside the incubator, sterilise
your hands and wipe the plate/dish with 70% ethanol
No metal racks to be put inside the incubator as these will
rust
Label all your work with your name/date/cell type
Report any sounding alarms (low CO2/humidity/water)
immediately to Helen (or Tony if Helen is absent)
Contamination of any plates/dishes should also be
immediately reported to Helen (or Tony if Helen is
absent) to prevent spread to other users’ work
Use of microscope

Please turn off microscope after use and cover it with the
proper microscope cover

If the microscope is damaged or not working, please
immediately report to Helen (or Tony if Helen is absent)
De-contamination of contaminated cultures

Any contaminated flasks/plates must be discarded (if the
user is absent, notify them first before discarding)

Contaminated plates must be bleached and rinsed before
discard
Accidents

Get first aid if needed and report immediately to Helen
(or Stephen/Tony if Helen is absent)
The following is for Helen to go through with the users:
-
-
-
All users must be ticked off by Helen to show they are competent at using the
facilities (use of the class II hood, waste disposal (liquid, solids, glass, chemicals)
Only tissue culture can be performed in the tissue culture laboratory. There are to
be absolutely NO animal dissections in the tissue culture room. Any animal
dissections should be carried out in room L9-15
Use your own reagents – do not use others’ reagents unless you ASK THEM FIRST!
If you make a mess – clean it up! The tissue culture room runs smoothly based on an
honesty system. To leave the TC room in a mess is really selfish and inconsiderate of
other users.
Users who do not comply with the rules will be banned in the future from using TC
facilities.
Use of the Class II hood
1) All usage must be logged in the logbook
2) Open the front panel and turn on the hood air flow and light
3) Let the air flow stabilise
4) Use 70% ethanol to sterilise the hood prior to use, as well as any other
equipment to be used inside the hood (ie: tip boxes, pipettes)
5) Dispose of all tips/glass pipettes into the sharps box, and all liquid waste into the
container containing bleach solution, contaminated waste into the autoclave bin
and non-contaminated waste to the normal bin (see waste disposal below for
more details)
6) After use of the hood, use 70% ethanol to wipe the used equipment as well as
the inside of the hood. This includes the clear front panel and the sides of the
hood. All spills must be cleaned up
7) Turn off the hood and wait for the air flow to fully stop before closing the clear
front panel
8) Suction 5-10mL bleach and check the liquid waste to ensure that it is empty. If it
is full, empty the waste into the sink with copious amounts of water. Add 10mL
bleach to the bottle before putting it back into its place
Contamination and de-contamination
1) Any contaminated flasks/plates must be discarded (if the user is absent, notify
them first before discarding
2) Contaminated plates must be bleached and rinsed before discard.
-
Waste disposal
Glass pipettes/sharps/pipette tips: to be discarded into the sharps container
Liquid waste: TC waste to be bleached and discarded into the sink with lots of water.
Solvents must be put into halogenated or non-halogenated waste bin in the main lab
(L9-14). Trizol to be put into special trizol waste. Solvents and other toxic chemicals
-
MUST NOT be poured down the sink. If you are unsure where to dispose something,
please ask Helen (or Tony if Helen is absent)
Solid TC waste: contaminated waste/tissue culture plates/dishes to be discarded into
the autoclave waste bin. No liquids are to be put here
Non-contaminated waste: to be discarded in normal bin
Use of the CO2 incubator
1) Ensure you use 70% ethanol to sterilise your hands prior to taking out your cell
culture
2) Prior to putting your cultures inside the hood, ensure you sterilise your hands
and wipe the plate/dish with 70% ethanol
3) No metal racks to be put inside the incubator as these will rust
4) All work must be labelled with your name/date/cell type
5) Any sounding alarms (low CO2/humidity/water) should be immediately reported
to Helen (or Tony if Helen is absent)
6) Contamination of any plates/dishes should also be immediately reported to
Helen (or Tony if Helen is absent) to prevent spread to other users’ work
Use of microscope
1) Please turn off microscope after use and cover it with the proper microscope
cover
2) If the microscope is damaged or not working, please immediately report to Helen
Accidents
1) If you should be unfortunate to be involved in a lab accident, ensure you and/or
the other party gets immediate first aid where applicable
2) Report all accidents to Helen (or Stephen or Tony if Helen is absent)
Summary checklist for Helen:
-
-
-
-
Booking in the logbook
Use of Class II hood
o Turning on/off and time for the air flow to stabilise
o Cleaning up spills/mess inside AND outside of the hood
Waste disposal
o TC plates
o TC media
o Solvents
o Non-contaminated dry waste
o Trizol
o Glass/pipette tips
Use of the CO2 incubator
o Procedure to put things in and take things out
o What do to if there is an alarm
Use of the microscope
What to do if an accident occurs
TC user test
1)
What should you wipe the Class II hood with before and after use?
2)
The Class II hood should be run for at least ____ minutes before use
3) Write down the correct method of disposal of the following items
- tissue culture media waste
_________________________________
- used tissue culture plates
_________________________________
- paper towels
_________________________________
- glass pipettes
_________________________________
- yellow tips
_________________________________
- 100% ethanol
_________________________________
- trizol
_________________________________
- chlorinated waste
_________________________________
- plastic pipettes
_________________________________
4)
What do you do if the CO2 alarm of the incubator is on?
5)
What should you do if you discover your TC plates are contaminated?
6)
What should you do with bleached plates/flasks before discarding them?
7)
True or false: A user may use the TC room for animal dissection ___________
8)
True or false: I can grow bacterial cultures in the CO2 incubator ___________
9)
True or false: I can tip anything down the sink as long as I flush it with lots of water
____________
10) True or false: My gloved hands and tissue culture plates/flasks must be wiped with
ethanol before putting plates/flasks back in the CO2 incubator ______________
By completing and passing this test, I understand that I will adhere to the rules of the
TC lab, and that if I do not follow the guidelines, this will result in my suspension
from use of TC facilities
Name:________________________
Signature:______________________
Date:_________________________
Authorised by:_________________