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This article was published in 1956
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INSTITUTE OF INSPECTORS OF STOCK OF N.S.W. YEAR BOOK.
VACCINE MANUFACTURE
Recent Developments
A. F. WEBSTER, B.V.Sc., Windsor Road, Northmead.
Some of the difficulties to be overcome in the preparation of vaccines commercially may be illustrated by reference to Leptospira pomona; the organism most commonly associated in Australia with outbreaks of Leptospirosis, a disease for which vaccine therapy should be most helpful. However, this organism is very sensitive to changes in culture media, even from batch to batch of the same formula, and is prone to dry out in stock culture. The habituation of outbreak strains to artificial culture is not achieved at all readily, and without special precautions the culture decreases in virulence and loses its power to promote temperature reactions in animals. In the absence of such reaction to test the efficiency of a vaccine it is not possible to assess the value of any particular batch of the product.
The growing of bacteria in semi-permeable tubes is one method which has been developed recently. Large cellophane tubes about 6 inches in diameter and 9 feet long are intussuscepted to give a double-walled tube, and then are suspended in the culture medium. This is held in a large pyrex container; shaped like a test tube, approximately 5 feet high and protected by a metal cage. The inside of the cellophane bag is filled with saline and the whole sterilized in an autoclave.
This method is used mainly when dealing with anaerobes; particularly Cl. chauvoei or Cl.septicus, for the production of Blackleg vaccine. When such anaerobes are to be grown, as soon as the medium is sufficiently cool after sterilisation, the seed culture is added through a long tube into the saline solution, and at the bottom of the bag. The nutriments of smaller molecular size pass through the bag wall and are available for use by the bacteria. It is probable that some of the growth products pass out, but the large molecule toxins and the bacterial bodies are held; giving finally a much stronger concentration of bacteria and toxins than can be achieved in free growth in broth. The end product is almost creamy in consistency, and it has been shown in the case of Cl. chauvoei that one-fiftieth ml. will give adequate immunity to calves and sheep. This means that the dose for use in the field, now generally 5 ml., could be reduced to any desired size. However, unexpected but unanimous opposition, by those engaged in distribution in the field, to any reduction in dose size has held up any alteration.
One advantage of this method of manufacture is that greater antigenic value can be concentrated in the 5 ml. dose, while a mixed chauvoei and septicus vaccine can be made without the content of either antigenic component being reduced. It has been stated also that the exclusion of a considerable proportion of the protein constituents of the culture medium from the vaccine leads to less distraction of the immune mechanism from the true disease antigens; the administration of which is the real purpose of vaccination.
Perhaps the most striking advance in the manufacturing field in recent years is the freeze-drying of biological products on a commercial scale. The first type used by the writer was a small cylinder containing a chemical drying agent; this cylinder being fitted with some forty nipples upon which ampoules could be plugged. A vacuum was drawn in the cylinder sufficient to boil and freeze the water contained in the ampoules. This method is suitable only for very small quantities, such as the preservation of stock cultures.
The first commercial freeze-dried vaccine was produced on an extension of this idea. A manifold holding some three hundred ampoules led into a chamber refrigerated at -50 degrees C. to trap the water removed and prevent it reaching the vacuum pump. A vacuum of about 0.1 mm. of Mercury was drawn on the system and the ampoules containing 2 ml. of liquid vaccine were spun mechanically in a freezing bath at -30 degrees to 40 degrees C. to give a frozen cylinder of greatest possible area on the inside of the ampoule. Immediately the ampoule was plugged into the manifold, the evaporation was sufficient to maintain the frozen state. In about one hour the tubes appear to be dry, but the process is continued for some 5-6 hours to remove all but a trace of moisture. The ampoules are then removed, the necks drawn out to facilitate sealing, replaced on the manifold and the vacuum again drawn. The ampoules are then sealed off under vacuum; being tested for leaks with a high frequency spark.
The drawbacks of this method are the fragility of the finished pack, the time consumed in the various manipulations and the lack of control of the temperature during drying. This latter is a major factor when the method is used to make dried Strain 19 Vaccine; in which, unless the temperature is controlled strictly throughout, the loss of organisms in drying reached 99% instead of about 40%.
The most recent installation is of the cabinet type in which the material to be dried is placed in bottles which eventually are sealed with rubber and aluminium caps. The four upper shelves of the cabinet are fitted with double coils so that they can either be cooled to about -10 degrees C. or heated to any desired temperture. The two lower shelves are cooled to 50 degrees C(sic). and serve as condenser plates; freezing out the water vapour and preventing it reaching the vacuum pumps.
The bottles containing the material to be dried are placed on the shelves and frozen. When the desired temperature is reached a vacuum is drawn in the cabinet and heat applied to the shelves supporting the bottles. This heat is regulated to keep the contents at the correct temperaure. When drying is completed the bottles are withdrawn and the caps applied but not sealed. They are replaced in the chamber and a vacuum again drawn to remove air and any moisture picked up from the atmosphere. This vacuum is then broken with pure dry nitrogen; after which the bottles are removed and sealed as quickly as possible with an automatic machine handling 2800 per hour.
The cabinet has a capacity of some 7000 single dose bottles of Distemper Vaccine and with this apparatus it is hoped to manufacture products not as yet available in Australia.