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Federal Register / Vol. 46, No. 233 / Friday, December 4, 1981 / Notices 
the environment. Special laboratory 
design is used primarily in facilities in 
which experiments of moderate to high 
potential hazards are performed. 
Combinations of laboratory practices, 
containment equipment, and special 
laboratory design can be made to 
achieve different levels of physical 
containment. Four levels of physical 
containment, which are designated as 
Pi, P2, P3, and P4. eire described. It 
should be emphasized that the 
descriptions and assignments of 
physical containment detailed below are 
based on existing approaches to 
containment of pathogenic organisms. 
For example, the “Classification of 
Etiologic Agents on the Basis of 
Hazard,"(7j prepared by the Centers for 
Disease Control, describes four general 
levels which roughly correspond to our 
descriptions for Pi, P2, P3, and P4; and 
the National Cancer Institute describes 
three levels for research on oncogenic 
viruses which roughly correspond to our 
P2. P3, and P4 levels.[6] 
It is recognized that several different 
combinations of laboratory practices, 
containment equipment, and special 
laboratory design may be appropriate 
for containment of specific research 
activities. The Guidelines, therefore, 
allow alternative selections of primary 
containment equipment within facilities 
that have been designed to provide P3 
and P4 levels of physical containment. 
The selection of alternative methods of 
primary containment is dependent, 
however, on the level of biological 
containment provided by the host-vector 
system used in the experiment. 
Consideration will also be given by the 
Director. NIH, with the advice of the 
Recombinant DNA Advisory Committee 
to other combinations which achieve an 
equivalent level of containment. (See 
Section IV-E-l-b-{2Hb).) 
Information on large-scale 
applications can be found in “Physical 
Containment Recommendations for 
Large-Scale Uses of Organisms 
Containing Recombinant DNA 
Molecules.” Federal Register, April 11, 
1980. where definitions are found of Pl- 
LS, P2-LS. and P3-LS. Guidance is 
available from ORDA on physical 
containment levels when working with 
plant host-vector systems. 
II-B-1. Pi Level. 
Il-B-l-a. Laboratory Practices. 
II-B-l-a-(l). Laboratory doors shall 
be kept closed while experiments are in 
progress. 
n-B-l-a-{2). Work surfaces shall be 
decontaminated daily, and immediately 
following spills of organisms containing 
recombinant DNA molecules. 
D-B-l-a-{3). All biological wastes 
shall be decontaminated before 
disposal. Other contaminated materials, 
such as glassware, animal cages, and 
laboratory equipment, shall be 
decontaminated before washing, reuse, 
or disposal. 
II-B-l-a-(4). Mechanical pipetting 
devices shall be used; pipetting by 
mouth is prohibited. 
Il-B-l-a-{5). Eating, drinking, 
smoking, and storage of foods are not 
permitted in the laboratory area in 
which recombinant DNA materials are 
handled. 
U-B-l-a-{6). Persons shall wash their 
hands after handling organisms 
containing recombinant DNA molecules 
and when they leave the laboratory. 
II-B-l-a-(7). Care shall be taken in 
the conduct of all procedures to 
minimize the creation of aerosols. 
II-B-l-a-{8). Contaminated materials 
that are to be decontaminated at a site 
away from the laboratory shall be 
placed in a durable leak-proof container, 
which is closed before removal from the 
laboratory. 
II-B-l-a-{9). An insect and rodent 
control program shall be instituted. 
Il-B-l-a-(10). The use of laboratory 
gowns, coats, or uniforms is 
discretionary with the laboratory 
supervisor. 
il-B-l-a-(ll). Use of the hupodermic 
needle and syringe shall be avoided 
when alternative methods are available. 
ll-B-l-a-{12). The laboratory shall be 
kept neat and clean. 
lI-B-l-b. Containment Equipment. 
Special containment equipment is not 
required at the PI level. 
II-B-l-c. Special Laboratory Design. 
Special laboratory design is not required 
at the Pi level. 
II-B-2. P2 Level. 
II-B-2-a. Laboratory Practices. 
Il-B-2-a-(l). Laboratory doors shall 
be kept closed while experiments are in 
progress. 
Il-B-2-a(2). Work surfaces shall be 
decontaminated daily, and immediately 
following spills of organisms containing 
recombinant DNA molecules. 
Il-B-2-a(3). All laboratory wastes 
shall be steam-sterilized (autoclaved) 
before disposal. Other contaiminated 
materials such as glassware, animal 
cages, laboratory equipment, and 
radiocative wastes shall be 
decontaminated by a means 
demonstrated to be effective before 
washing, reuse, or disposal. 
II-B-2-a(4). Mechanical pipetting 
devices shall be used; pipetting by 
mouth is prohibited. 
lI-B-2-a(5). Eating, drinking, smoking, 
and storage of food are not permitted in 
the laboratory area in which 
recombinant DNA materials are 
handled. 
II-B-2-a(6). Persons shall wash their 
hands after handling organisms 
containing recombinant DNA molecules 
and when they leave the laboratory. 
lI-B-2-a(7). Care shall be exercised to 
minimize the creation of aerosols. For 
example, manipulations such as 
inserting a hot inoculating loop or 
needle into a culture, flaming an 
inoculation loop or needle so that it 
splatters, and forceful ejection of fluids 
from pipettes or syringes shall be 
avoided. 
lI-B-2-a(8). Contaminated materials 
that are to be steam sterilized 
(autoclaved) or decontaminated at a site 
away from the laboratory shall be 
placed in a durable leak-proof container, 
which is closed before removal from the 
laboratory. 
Il-B-2-a(9). Only persons who have 
been advised of the nature of the 
research being conducted shall enter the 
laboratory. 
lI-B-2-a(10). The universal biohazard 
sign shall be posted on all laboratory 
access doors when experiments 
requiring P2 containment are in 
progress. Freezers and refrigerators or 
other units used to store organisms 
containing recombinant DNA molecules 
shall also be posted with the universal 
biohazard sign. 
II-B-2-a(ll). An insect and rodent 
control program shall be instituted. 
II-B-2-a(12). The use of laboratory 
gowns, coats, or uniforms is required. 
Laboratory clothing shall not be worn to 
the lunch room or outside of the building 
in which the laboratory is located. 
II-B-2-a(13). Animals not related to 
the experiment shall not be permitted in 
the laboratory. 
Il-B-2-a(14). Use of the hypodermic 
needle and syringe shall be avoided 
when alternative methods are available. 
II-B-2-a(15). The laboratory shall be 
kept neat and clean. 
II-B-2-a(16). Experiments of lesser 
biohazard potential can be carried out 
concurrently in carefully demarcated 
areas of the same laboratory. 
II-B-2-b. Containment Equipment. 
Biological safety cabinets [20] shall be 
used to contain aerosol-producing 
equipment, such as blenders, 
lyophilizers, sonicators, and centrifuges, 
when used to process organisms 
containing recombinant DNA molecules, 
except where equipment design 
provides for containment of the 
potential aerosol. For example, a 
centrifuge may be operated in the open 
if a sealed head or safety centrifuge 
cups are used. 
I1-B-2-C. Special Laboratory Design. 
An autoclave for sterilization of wastes 
and contaminated materials shall be 
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