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Federal Register / Vol. 49, No. 227 / Friday, November 23, 1984 / Notices 
Appendix K-JV-C. Sample collection 
from a closed system, the addition of 
materials to a closed system, and the 
transfer of culture fluids from one closed 
system to another shall be done in a 
manner which prevents the release of 
aerosols or contamination of exposed 
I surfaces. 
Appendix K-IV-D. Exhaust gases 
removed from a closed system or other 
primary containment equipment shall be 
treated by filters which have efficiencies 
equivalent to HEPA filters or by other 
equivalent procedures (e.g., incineration 
to prevent the release of viable 
organisms containing recombinant DNA 
molecules to the environment. 
Appendix K-IV-K A closed system or 
other primary containment equipment 
that has contained viable organisms 
containing recombinant DNA molecules 
shall not be opened for maintenance or 
other purposes unless it has been 
sterilized by a validated sterilization 
procedure. A validated sterilization 
; procedure is one which has been 
demonstrated to be effective using the 
organisms that will serve as the host for 
propagating the recombinant DNA 
i molecules. 
Appendix K-IV-F. A closed system 
used for the propagation and growth of 
viable organisms containing 
recombinant DNA molecules shall be 
operated so that the space above the 
culture level will be maintained at a 
pressure as low as possible, consistent 
| with equipment design, in order to 
maintain the integrity of containment 
features. 
Appendix K-IV-G. Rotating seals and 
other mechanical devices directly 
associated with a closed system used to 
contain viable organisms containing 
recombinant DNA molecules shall be 
designed to prevent leakage or shall be 
fully enclosed in ventilated housings 
that are exhausted through filters which 
have efficiencies equivalent to HEPA 
filters or through other equivalent 
treatment devices. 
Appendix K-IV-H. A closed system 
used for the propagation and growth of 
viable organisms containing 
recombinant DNA molecules and other 
primary containment equipment used to 
contain operations involving viable 
organisms containing recombinant DNA 
molecules shall include monitoring or 
sensing devices that monitor the 
integrity of containment during 
operations. 
Appendix K-IV-I. A closed system 
I used for the propagation and growth of 
viable organisms containing 
recombinant DNA molecules shall be 
tested for integrity of the containment 
features using the organisms that will 
serve as the host for propagating the 
recombinant DNA molecules. Testing 
shall be accomplished prior to the 
introduction of viable organisms 
containing recombinant DNA molecules 
and following modification or 
replacement of essential containment 
features. Procedures and methods used 
in the testing shall be appropriate for the 
equipment design and for recovery and 
demonstration of the test organism. 
Records of tests and results shall be 
maintained on file. 
Appendix K-IV-J. A closed system 
used for the propagation and growth of 
viable organisms containing 
recombinant DNA molecules shall be 
permanently identified. This 
identification shall be used in all records 
reflecting testing, operation, and 
maintenance and in all documentation 
relating to the use of this equipment for 
research production activities involving 
viable organisms containing 
recombinant DNA molecules. 
Appendix K-IV-K. The universal 
biohazard sign shall be posted on each 
dosed system and primary containment 
equipment when U9ed to contain viable 
organisms containing recombinant DNA 
molecules. 
Appendix K-IV-L Emergency plans 
required by Section IV-B-3-f shall 
include methods and procedures for 
handling large losses of culture on an 
emergency basis. 
Appendix K-IV-M. Closed systems 
and other primary containment 
equipment used in handling cultures of 
viable organisms containing 
recombinant DNA molecules shall be 
located within a controlled area which 
meets the following requirements: 
Appendix K-IV-M-1. The controlled 
area shall have a separate entry area. 
The entry area shall be a double-doored 
space such as an air lock, anteroom, or 
change room that separates the 
controlled area from the balance of the 
facility. 
Appendix K-IV-M-2. The surfaces of 
walls, ceilings, and floors in the 
controlled area shall be such as to 
permit ready cleaning and 
decontamination. 
Appendix K-IV-M-3. Penetrations 
into controlled area shall be sealed to 
permit liquid or vapor phase space 
decontamination. 
Appendix K-fV-M-4. All utilities and 
service or process piping and wiring 
entering the controlled area shall be 
protected against contamination. 
Appendix K-IV-M-5. Hand-washing 
facilities equipped with foot, elbow, or 
automatically operated valves shall be 
located at each major work area and 
near each primary exit. 
Appendix K-IV-M-6. A shower 
facility shall be provided. This facility 
shall be located in close proximity to the 
controlled area. 
Appendix K-1V-M-7. The controlled 
area shall be designed to preclude 
release of culture fluids outside the 
controlled area in the event of an 
accidental spill or release from the 
closed systems or other primary 
containment equipment. 
Appendix K-IV-M-8. The controlled 
area shall have a ventilation system that 
is capable of controlling air movement. 
The movement of air shall be from areas 
of lower contamination potential to 
areas of higher contamination potential. 
If the ventilation system provides 
positive pressure supply air, the system 
shall operate in a manner that prevents 
the reversal of the direction of air 
movement or shall be equipped with an 
alarm that would be actuated in the 
event that reversal in the direction of air 
movement were to occur. The exhaust 
air from the controlled area shall not be 
recirculated to other areas of the 
facility. The exhaust air from the 
controlled area may be discharged to 
the outdoors without filtration or other 
means for effectively reducing an 
accidental aerosol burden provided that 
it can dispersed clear of occupied 
buildings and air intakes. 
Appendix K-IV-M-. The following 
personnel and operational practices 
shaH be required: 
Appendix K-lV-M-1. Personnel entry 
into the controlled area shall be through 
the entry area specified in Appendix K- 
1V-M-1. 
Appendix K-IV-M-2. Persons entering 
the controlled area shall exchange or 
cover their personal clothing with work 
garments such as jumpsuits, laboratory 
coats, pants and shirts, hear cover, and 
shoes or shoe covers. On exit from the 
controlled area the work clothing may 
be stored in a locker separate from that 
used for personal clothing or discarded 
for laundering. Clothing shall be 
decontaminated before laundering. 
Appendix K-IV-N-3. Entry into the 
controlled area during periods when 
work is in progress shall be restricted to 
those persons required to meet program 
or support needs. Prior to entry all 
persons shall be informed of the 
operating practices, emergency 
procedures, and the nature of the work 
conducted. 
Appendix K-IV-N-4. Persons under 18 
years shall not be permitted to enter the 
controlled area. 
Appendix K-IV-M-5. The universal 
biohazard sign shall be posted on entry 
doors to the controlled area and all 
internal doors when any work involving 
the organism is in progress. This 
includes periods when decontamination 
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