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Federal Register / Vol. 46, No. 126 / Wednesday. July 1, 1981 / Notices 
that investigators and institutional 
biosafety committees recommend 
changes in the Guidelines to permit their 
use. 
II-A. Standard Practices and 
Training. The first principle of 
containment is a strict adherence to 
good microbiological practices. [6-15] 
Consequently, all personnel directly or 
indirectly involved in experiments on 
recombinant DNAs must receive 
adequate instruction, (see Sections IV- 
D-l-g, IV-D-5-d and IV-D-8-b.). This 
shall, as a minimum, include instructions 
in aseptic techniques and in the biology 
of the organisms used in the 
experiments, so that the potential 
biohazards can be understood and 
appreciated. 
Any research group working with 
agents with a known or potential 
biohazard shall have an emergency plan 
which describes the procedures to be 
followed if an accident contaminates 
personnel or the environment. The 
principal investigator must ensure that 
everyone in the laboratory is familiar 
with both the potential hazards of the 
work and the emergency plan. (See 
Sections IV-D-5-e and IV-D-3-d.) If a 
research group is working with a known 
pathogen where there is an effective 
vaccine it should be made available to 
all workers. Where serological 
monitoring is clearly appropriate it shall 
be provided. (See Sections IV-D-l-h 
and IV-D-8-C.) 
II-B Physical Containment Levels. 
The objective of physical containment is 
to confine organisms containing 
recombinant DNA molecules, and thus 
to reduce the potential for exposure of 
the laboratory worker, persons outside 
of the laboratory, and the environment 
to organisms containing recombinant 
DNA molecules. Physical containment is 
achieved through the use of labor^'tory 
practices, containment equipment, and 
special laboratory design. Emphasis is 
placed on primary means of physical 
containment which are provided by 
laboratory practices and containment 
equipment. Special laboratory design 
provides a secondary means of 
protection against the accidental release 
of organisms outside the laboratory or to 
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, are 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,” (7) 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. (5) 
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 the 
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-(2)-(b).) Additional 
material on physical containment for 
plant host-vector systems is found in 
Sections III-C-3 and III-C-4. 
II-B-1. Pi Level, 
II-B-l-a. Laboratory Practices. 
II-B-l-a-(l). Laboratory doors shall 
be kept closed while experiments are in 
progress. 
II-B-l-a-(2j. Work surfaces shall be 
decontaminated daily, and immediately 
following spills of organisms containing 
recombinant DNA molecules. 
II-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-^l-a-(4). Mechanical pipetting 
devices shall be used; pipetting by 
mouth is prohibited. 
II-B-l-a-(5j. Eating, drinking, 
smoking, and storage of foods are not 
permitted in the laboratory area in 
which recombinant DNA materials are 
handled. 
II-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. 
Il-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. 
II-B-l-a-(lO). The use of laboratory 
gowns, coats, or uniforms is 
discretionary with the laboratory 
supervisor. 
II-B-l-a-(ll). Use of the hypodermic 
needle and syringe shall be avoided 
when alternative methods are available. 
U-B-l-a-(12). The laboratory shall be 
kept neat and clean. 
II-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. 
II-B-2-a-(l). Laboratory doors shall 
be kept closed while experiments are in 
progress. 
II-B-2-a-(2). Work surfaces shall be 
decontaminated daily, and immediately 
following spills of organisms containing 
recombinant DNA molecules. 
II-B-2-a-(3). All laboratory wastes 
shall be steam-sterilized (autoclaved) 
before disposal. Other contaminated 
materials such as glassware, animal 
cages, laboratory equipment, and 
radioactive wastes shall be 
decontaminated by a means 
demonstrated to be effective before 
washing, reuse, or disposal. 
n-B-2-a-(4). Mechanical pipetting 
devices shall be used; pipetting by 
mouth is prohibited. 
II-B-2-a-(5). Eating, drinking, 
smoking, and storage of food are not 
permitted in the laboratory area in i 
which recombinant DNA materials are 
handled. j 
II-B-2-a-(6). Persons shall wash their , 
hands after handling organisms . 
containing recombinant DNA molecules ' 
and when they leave the laboratory. 
II-B-2-a-(7). Care shall be exercised | 
to minimize the creation of aerosols. For i\ 
example, manipulations such as < 
inserting a hot inoculating loop or ' 
needle into a culture, flaming an i 
inoculation loop or needle so that it ,( 
splatters, and forceful ejection of fluids 
from pipettes or syringes shall be 
avoided. t 
II-B-2-a-(8). Contaminated materials 
that are to be steam sterilized Ij 
(autoclaved) or decontaminated at a site 3 
away from the laboratory shall be ? 
placed in a durable leak-proof container li 
[152] 
