[NATIONAL LIBRARY OF MEDICINE FREDERICK FACILITY HF3412 16 mm COLOR PRINT OPTICAL SOUND]

[THE P4 CONTAINMENT FACILITY FOR RECOMBINANT DNA RESEARCH]

[Located at THE FREDERICK CANCER RESEARCH CENTER Frederick, Md.]

[NATIONAL INSTITUTES OF HEALTH]

[Narrator:] The first certified P4 containment facility in the United States for recombinant DNA research,

a laboratory designed to meet the requirements set forth in the NIH guidelines governing recombinant DNA research,

is now available to scientists.

The laboratory itself is under negative air pressure.

The experiments are carried out from beginning to end inside an elaborate system of enclosed gas-tight cabinets.

This means that nothing, not even the tiniest particle of dust or air,

or the smallest micro-organism,

can penetrate the heavy stainless steel and glass cabinets.

This system of cabinets measures about seventy feet in combined length.

It is contained inside a room which has an area of about four hundred square feet.

Using the shoulder-length rubber gloves, which are tightly attached to the sides of the cabinet,

laboratory workers are able to move and control all the materials and tools used in the experiment without direct contact

between their hands and anything inside the cabinet.

Each experiment must be carefully planned well in advance, since all of the materials used in the experiment,

including chemicals, cultures, and laboratory equipment, must be brought 

into the group of interconnected cabinets, or cabinet line, as it is called.

One of the methods of bringing materials into the cabinet is through an autoclave, or steam sterilizer.

Equipment and materials are loaded onto a shelf inside the autoclave for transfer to the interior of the cabinet lines.

A heavy door is then swung into place and locked to provide a tight seal.

A laboratory worker removes them from the cabinet side of the autoclave. 

Because the interior of the device has now been exposed

to the inside of the cabinet,

it will be necessary to run the autoclave

through a sterilizing cycle before it is opened again on its outside end.

The two-hundred and fifty degree Fahrenheit steam pressure cycle takes time. 

All large items have to enter through the autoclave, so planning its maximum use is essential.

The cabinet lines are modular, that is,

made up of individual connected units allowing the free movement of materials within the lines.

At least two workers are required for most manipulations and movements of materials.

A variety of equipment for experiments must be contained within the cabinets,

and that includes a refrigerator which is used for both long- and short-term storage of materials.

Here, supplies of growth media are being checked and readied for later use in preparing bacterial culture.

Another designed feature of the refrigerator is movable shelves which slide in and out

to allow the laboratory worker easier access to materials stored within.

Note the tools used to extend the limited reach of the researcher

as he works within the restricted space of the gas-tight cabinets.

The cabinet system in this laboratory has two levels which are connected by a small elevator or dumbwaiter.

Specially weakened strains of bacteria

which cannot survive outside of the carefully controlled laboratory conditions,

are grown in a waterbath located on the second tier of the system. 

Once the cultures have been grown and are ready for use in the experiment,

they are transferred to the nearby elevator,

which descends with them to the lower level.

Here, most of the biochemical and microbiological steps in the experiment are actually carried out.

The weakened strains of bacteria which are used in recombinant DNA research

make up a second form of containment called biological containment.

[simulated test]

Although physical barriers or physical containment

are highly effective in preventing the escape of micro-organisms,

biological containment adds a built-in safeguard.

The crippled bacteria are so weak that they self-destruct outside the laboratory.

These two types of barriers, physical and biological, are always used in combination.

This particular cabinet system has an area with glove ports on both sides.

This arrangement allows two laboratory workers to team up

for some of the more delicate operations,

such as the transfer of cultures or reagents from one container to another.

Continuing the inventory of research tools mounted within the cabinet line is a high speed centrifuge.

This device, mounted flush with the bottom space of the cabinet,

is using in purifying DNA.

Spinning rapidly, DNA separates from protein and solvent. 

The control panel for this centrifuge is mounted outside the barrier of the gas-tight cabinet.

Bacteria for experiments such as this are grown in single colonies which are cultivated in standard laboratory petri dishes.

The petri dishes are kept in an incubator inside the cabinet line.

The microscope is a vital tool in experiments involving micro-organisms.

An instrument is mounted in this laboratory's line.

Despite the restraints that are imposed by the heavy rubber gloves,

skilled laboratory workers are able to make delicate adjustments on the microscope.

A closed-circuit television system with the screen mounted just above the researcher's head provides the means for viewing.

A second means of entering the enclosed cabinet line is through a device called a dunk tank.

This consists of a tank filled with a strong liquid disinfectant and is an integral part of the barrier,

and like the autoclave, prevents the passage of unwanted living organisms in either direction.

A leakproof container is used to convey materials needed inside the line, safely through the dunk tank.

Note how the attached gloves are pulled back inside the cabinet.

This is due to the negative air pressure that is constantly maintained within the cabinets.

This added safety feature ensures that no organism escapes, should the gloves be punctured.

Although use of experimental animals is not a significant part of most recombinant DNA research,

this laboratory is equipped to house animals on two tiers within the cabinets.

Experimental mice like these can be inoculated and examined within the cabinets.

The daily care and feeding of the animals is also carried out entirely within the cabinet system.

Materials needed to work with and care for the animals are introduced through the dunk tank.

When necessary, cages and waste are removed in the same fashion that all other material is taken out: 

through an autoclave.

The material is placed inside and is then heated to a temperature sufficient to kill all organisms.

At the end of a sterilization cycle,

material is removed from the autoclave for further processing,

either washing if it is reusable laboratory equipment,

or disposal as trash if it is not reusable.

As any material leaves the building, it is sterilized a second time.

Material is loaded into the autoclave and again heated to achieve complete sterilization.

[A Closer Look at DNA]

We have seen the facilities, equipment, and techniques that will be used in this laboratory.

Now let's take a close look at the substance

which is the focus of the scientific activity which will be carried out here.

In order to show more clearly some of the methods of DNA research,

we are going through a non-recombinant DNA procedure.

Close-up pictures, which you will see,

of a bacterial DNA culture were made under P3 conditions.

Reagents are added to the purified DNA to separate it from other materials

such as protein and to prepare it for spinning.

A glass rod can then be used to spin the purified DNA into a cohesive mass.

Pressed against the side of the test tube to eliminate excess moisture,

the long strands of DNA take on the appearance of transparent thread,

wrapped around the end of the glass rod.

The DNA molecules may be dissolved again in a salt solution

for further study and analysis of the DNA's genetic or biochemical make-up if so desired.

At this stage the DNA is ready for spinning or other processing as may be be called for in the experiment.

[Film ends]