1. When transporting anything you need to bring from upstairs – place inside the sterile container (with the blue handle). All materials, placed in the box, need to be wiped with Bacillol.

2. After entering the room, turn on the light and the switch above – airflow.

3. Take off all the bracelets or watches, wash hands up to the elbows, disinfect hands and let the disinfectant dry.

4. Use only the lab coat, dedicated for the tissue culture. Keep it in the room. You’re always welcome to use my coat.

5. Everything in the room should be touched only with gloves.

6. Make sure that the pump waste bottle is less than not too full ¾ full (the line shows the maximum filling point)

7. Turning on the hood:

• press the button on the top for approx. 3 sec
• on the screen inside the hood the green and red lights should turn on. Don’t rise the glass yet
• turn on the UV by pressing “DIS” button on the remote control (press long until the UV lights up)
• UV the hood for 0,5-1h, there is the timer on the screen
• press “DIS” button again to switch off the UV
• open the hood with the button on the top
• the glass needs to go to the level where the double-arrows light turns green
• after a few seconds the airflow light will turn green as well

8. The hood is ready to operate

9. When starting to work under the hood – change to the new pair of gloves and use the plastic sleeves (bottom drawer under the sink) to cover the skin between the lab coat and gloves.

10. Wet the surface of the hood with the 75-80% EtOH, let it stay for approx.30 sec. Wipe the EtOH.

11. Wipe hands and sleeves with EtOH every time, when going under the hood and if you stayed outside the hood for more than 5 sec

12. Take your time to arrange everything under the hood from dirty to clean (usually left to right).

13. Don’t do quick movements under the hood (they can disturb the air flow and reduce the sterility). Put everything you need under the hood in advance. Minimize going in and out of the hood.

14. Everything what goes under the hood needs to be wiped with EtOH.

15. Turn on the pump. Clean the pump tubing with EtOH.

16. Even under the hood, minimize the time for any solution or cell culture plate to stay open.

17. Don’t touch:

• anything with the plastic or glass pipettes after you opened them or took out from the metal box
• rims of falcons when aspirating something
• other glass pipettes in the box, except for the one you are taking out

18. Do only sterile pouring under the hood to not create aerosols.

19. Trash the serological plastic pipettes (metal bin) separately from wrappers (floor and when you’re done – plastic bin). Glass pipettes go to the black square container under the hood

20. Make aliquotes for everything you use. Mark aliquotes with the date, when you start using it, not just when you aliquoted it.

21. After you’re done – clean the hood again. Clean the pump tubing with EtOH.

22. Switch the hood off by pressing the button on the top, bring the glass down and wait for approx. 5 sec for all the lights on the screen of the hood to turn off.

23. Switch off:

• hood
• pump
• microscopes (and cover them)
• centrifuge
• light and ventilation

24. Beads bath stays always on

If you're doing the cell culture for the first times - please read the info below.

Please find the more general rules for sterile work here: extended sterile work guidelines

The incubator model is HERAcell Vios 160i

1. Switch the incubator off (left side of the incubator, close to the bottom).

2. Take out the metal+plastic grid from the bottom front part and the filter from the black plastic “vacuum looking” part in the back. If you're sterilising several incubators at the same time, put these parts in separate plastic bags to not mix them up. The “vacuum looking” part can be placed back without the filter.

3. Wipe with EtOH all the shelves and the glass door.

4. Close the glass door and the main door, turn on the incubator.

5. Start the sterilisation on the screen, follow the instructions.

6. Open both doors and wait until the incubator beeps once (circa 30-60 sec).

7. Close the door, the procedure will take around 24h.