PC3 Laboratories

Two PC3 laboratories are located at SASVRC, Building 65, Royal Children’s Hospital in Rooms 518 and 607, on the 5th and 6th floors respectively. Access to these containment laboratories is confined to registered users who will undertake only approved work.
Entry is via an airlock with a key obtained from the administrative office on the 7th floor. The key must be signed out in the log book. Registered users must not allow non-registered personnel to use the key, nor allow them entry to the PC3 laboratories.

NOTE: Work with Risk Group 4 pathogens (as stated in AS 2243.3) or PC4 level recombinant DNA is not permitted under any circumstances.

The control and management of the PC3 Laboratories and the nature of the work performed in them is the responsibility of theDirector, SASVRC. Day-to-day management of individual laboratories is the responsibility of the Unit Head in charge. Administrative functions associated with the laboratories have been delegated to the SASVRC Safety Officer (hereafter “Safety Officer”). Conduct of PC3 recombinant DNA experimentation requires submission to the University of Queensland Institutional Biosafety Committee (IBC) for approval by the OGTR prior to commencement. The use of PC3 level, non-recombinant infectious agents or quantities (>500ml) of PC2 level, non-recombinant infectious agents designated as PC3 hazard, requires the written approval of the IBC prior to commencement.
All work conducted in the PC3 laboratories must comply with:

Users of the PC3 laboratories must obtain the necessary approvals for work with:

Users of the PC3 laboratories must obtain the appropriate vaccination(s) as recommended.

NOTE: Immuno-suppressed or immuno-compromised personnel may not work in the laboratory.

The laboratory is ventilated by a full outside air conditioning system with a dedicated fan coil unit. The laboratory is maintained at a negative pressure relative to the airlock and the corridor. The exhaust air is filtered through a pre-filter and an absolute HEPA filter before being vented through a single duct system via a duct mounted axial fan in the Level 6 plant room. Filtered air is exhausted to the outside of the building via a stack on the roof.
The exhaust fan is programmed to maintain a negative room pressure of 50 Pascals (Pa) with both air lock doors closed. An air conditioning control panel is mounted on the wall outside of the laboratory and is fitted with the following:

A differential pressure gauge is installed adjacent to the primary filter to indicate the pressure drop across the filter. The filter should be replaced when the reading indicates 125 Pascals.

A wall mounted switch outside the laboratory will close the supply air and exhaust air volume control dampers and also shut down the exhaust and supply air system.

A small circulating fan and tubular duct are fitted at high level between the air lock and filter plenum to facilitate decontamination of the room filters.

Entry to the laboratory is via the airlock. Registered users should use their key for the outer door and enter the airlock. Close the door behind you. If the outer door is not closed, and you open the inner door, an alarm will sound ( lab 618 only).
Enter the laboratory, and close the inner door securely behind you.
To exit the laboratory, follow the reverse procedure. Turn the door release handle on the inner door and proceed to the airlock, closing the door behind you. Open the outer door by turning the door handle and proceed to the corridor, closing the outer door behind you.
Under no circumstances are maintenance or service personnel allowed to enter unless the surfaces in the room have been decontaminated and the safety officer notified. It is preferable that all maintenance and equipment service, including fire extinguishers, shall be done at the time of the annual decontamination

The ventilation system is monitored by two Magnehelic gauges outside the laboratory measuring the pressure drop across the pre-filter and the differential air pressure between outside and inside the Laboratory.

On entering the laboratory the pressure differential will drop to approximately negative 25 Pa and will rise to approximately negative 50 Pa soon after the inner door is closed.
In the case of a failure in the ventilation system (pressure differential drops below negative 25 Pa for 30 seconds) an audible alarm will sound (the negative pressure alarm). You should immediately read the appropriate gauge and, if the differential pressure is less than 50 Pa, make all scientific work safe and secure* and then leave the laboratory and report the failure to the Safety Officer immediately.
* This may include turning off any equipment such as centrifuges or Bunsen burners, putting lids on petri dish cultures or solutions etc.
If the pre-filter pressure drop increases above 125 Pa, report this immediately to the Safety Officer

Work in laboratories at PC3 level shall conform with the procedures set out for PC2 laboratories. In addition, the following requirements apply:

All culture manipulations must be undertaken in the class II biosafety cabinet, with gloves. All aerosol-producing equipment such as that for sonication and vortexing shall be kept and used in the biosafety cabinet.
The biosafety cabinet and/or the laboratory shall be decontamined with formaldehyde gas after major spills of contaminated material.
No other work is to be done simultaneously with work requiring PC3 containment.
While work is in progress a sign on the door shall indicate the level of containment required for that work.
Laboratory clothing shall be laundered regularly. Protective clothing shall not be worn outside the laboratory; it shall be transported to the 6th floor autoclave in sealed bags or boxes. Boxes shall have provision for penetration of steam during autoclaving. Personal clothing and effects shall be kept in storage facilities located adjacent to the laboratory area and shall not be taken into the laboratory. Laboratory gowns worn in pC2 laboratories must not be taken into PC3 laboratories.
All equipment specified for PC3 requirements shall be dedicated to that use and area.
The laboratory doors shall be locked when the room is unoccupied.
All liquid waste is to be placed in Shott-bottles. In turn, these are to be placed in yellow containers with lids before transport to the 6th floor autoclave. Label with your name, date and name of infectious agent.
All solid waste is to be double bagged in autoclave bags before transport to the 6th floor autoclave. The person conducting the project in the PC3 is responsible for autoclaving the autoclave.
All work carried out in the PC3 shall be recorded in the log book.
Projects involving vaccinia virus are carried out in the PC3. Therefore, all users of the PC3 are to be vaccinated. (ie. Including those not directly involved in vaccinia-related projects).
Certain in vitro assays conducted in the PC3, eg 51Cr-release assays generate liquid waste with potentially low levels of contamination with infectious organisms. This liquid waste is to be decontaminated with disinfectant. An equal volume of sodium hypochlorite 10% freshly diluted to 1% (=10,000 ppm available chlorine) is appropriate. For details refer to B.12, PC2 laboratories guidelines.
At the completion of each session of work, surfaces of benches , the biosafety cabinet and other equipment used must be cleaned and decontaminated by wiping down with a fresh solution of an appropriate disinfectant. It is the responsibility of the researcher to determine the most appropriate disinfectant for use in their work (see AS/NZS 2243.3 Appendix E). The choice of disinfectant must take into account not only the types of microorganisms present, but also the factors affecting activity of disinfectants (e.g. contact time, concentration, inactivation by organic matter) and any other properties (e.g. flammability, toxicity, sensitizing action). Paper towel etc used for cleaning and disinfection must be placed in the discard can for autoclaving.
Open cuts and wounds including dermatitis should be well covered before entering the room.
Transport between the facility and any other PC3 facility will be in accordance with current Transport regulations and GTR Guidelines.
For transport within the building cultures should be placed inside a snaplock bag, padded with enough absorbent material in case of leakage, and then placed in an outer sealed unbreakable container.
Transport to other institutions outside of the building should be done to IATA standard

All spills of cultures must be dealt with promptly according to the following procedures.

Any small spillage must be decontaminated immediately by mopping up in paper towel followed by surface decontamination with paper towel soaked in the appropriate disinfectant. Disposable gloves must be worn. All contaminated paper towel and gloves must be double bagged and discarded in the containers for autoclaving.

Contact details for Registered Users are displayed next to the telephone in each PC3 laboratory. Protective clothing must be worn during decontamination.

1. Any major spillage (>10 ml ) or where personal contamination or splashing of walls has occurred must be reported immediately to another Registered User and/or the Safety Officer by telephone..

Decontamination of surfaces should be conducted as for small spills. Where there is a situation of personal risk such as when aerosol has been created, the user should telephone another Registered User, and await further instructions. Do not open the outer door unless you are in personal danger, for example due to fire.

2. After decontamination, report the incident to the Safety Officer and complete a University of Queensland Injury, Illness & Incident Report Form. A follow up consultation with the University Health Service may be required.

Personal contamination – requires the assistance of other Registered Users

If users are personally contaminated the following procedure should be followed:

Report the incident to another Registered User by telephone and, provided you are not in personal danger, wait in the airlock for assistance to arrive.
Remove contaminated clothing and place in a biohazard bag for autoclaving.
Decontaminate yourself by swabbing with paper towel soaked with a solution of Hibiclens.
Another Registered User will assist by ensuring that decontamination is completed, providing a clean change of clothing and if necessary transferring the user to the emergency shower and eyebath located outside of the 6th floor PC3 laboratory (lab 618).
Do not leave the airlock before decontamination unless your personal safety is at risk due to fire etc.
Report the incident to the Safety Officer and complete a University of Queensland Injury, Illness & Incident Report Form. A follow up consultation with the University Health Service may be required.

The laboratory will be routinely decontaminated with formaldehyde gas on an annual basis and after major spills of genetically manipulated or infectious agents. Decontamination will be carried out by NATA certified operators. After decontamination, provision will be made for laboratory equipment to be serviced and any other necessary maintenance to be carried out before re-commencement of work.

Formaldehyde decontamination and testing of cabinets will be carried out annually by NATA certified operators in accordance with Australian Standard AS 2647 Appendix C. Formaldehyde decontamination will be carried out following major spills of genetically manipulated or infectious agents.