Types and preparation of vaccines, pharmaceutical biotechnology

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Contents
Topic Page No.
Defination - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 2
Types - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 2-3
Preparation - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 3-4
Impact of genetic engineering on vaccine technology - - - - - - - 5
Quality control - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 5-6
Storage - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 6
References - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 7

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Definition:
Vaccine (L. vacca = cow) is a preparation/suspension or extract of a dead/attenuated (weakened)
germs of a disease which on inoculation (injection) into a healthy person provides
temporary/permanent, active/passive immunity by inducing antibodies formation. Thus antibody
provoking agents are called vaccines. Vaccines may be prepared from one species only or from a
mixture of two or more species.
The process of introduction of vaccine into an individual to provide
protection against a disease is called vaccination. Vaccination and immunization are two
different processes. Vaccination only refers to the administration of a vaccine or toxoid, while
immunization is the process by which the body produces antibodies against the vaccine
preventable diseases through administration of specific vaccines.
Types:
Vaccines are of three types:
(i) Killed (inactivated) vaccines: consist of microorganisms killed by heat or chemicals.
(ii) Live attenuated vaccines: consist of live bacteria or viruses which have been rendered
avirulent. They nevertheless grow and multiply in the body of the host to a limited extent.
In individuals with impaired host defence, e.g.
(a) Leukaemia or other malignancies, especially those receiving cytotoxic
chemotherapy.
(b) Systemic lupus erythematosus.
(c) Corticosteroid recipients.
(d) AIDS and other immune deficiency states.
The limited virulence of organisms in the live
vaccine may be sufficient to cause a disease; live vaccines are contraindicated in them.
Two live vaccines, if not given together, should preferably be administered with a gap
of 1 month.

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(iii) Toxoids: are modified bacterial exotoxins so that toxicity is lost but antigenicity is
retained. The term 'vaccine' is sometimes restricted to preparations of whole
microorganisms and toxoids are enumerated separately.
Preparation:
Vaccines may be prepared by the general methods given below or in any other manner, provided
the identity of the antigens is maintained and the vaccines are free from microbial contamination
and extraneous agents. Suitable adjuvants may be added during the preparation but streptomycin,
penicillin or other β-lactam antibiotics may not be added at any stage of manufacture or in the final
vaccine. A suitable bactericide may be added to sterile and inactivated vaccines. The final products
are distributed aseptically into sterile containers which are then sealed to exclude extraneous
micro-organisms.
Bacterial Vaccines:
Bacterial vaccines are either sterile suspensions of live or killed bacteria or sterile
extracts of derivatives of bacteria. They may be simple vaccines prepared from one species or may
be mixed vaccines prepared by blending two or more simple vaccines from different species or
strains.
Bacterial vaccines may be prepared from cultures grown on suitable solid or liquid media.
The whole culture or parts of it may be used in preparing the vaccine. The identity, antigenic
potency and purity of each bacterial culture must be carefully controlled.
Vaccines containing killed organisms may be prepared by killing the
organisms by chemical or physical means provided the antigenic potency of the vaccine is
preserved. Vaccines containing living bacteria may be prepared from strains which are avirulent
for humans but which stimulate the production of antibodies active against pathogenic strains of
the same species. The final vaccines must be free from any substance known to cause toxic, allergic
or other undesirable immunological reactions in humans.
Bacterial vaccines are suspensions of varying degrees of opacity in colourless or
slightly coloured liquids or they may be freeze-dried so that the water content is not more than 3.0
per cent w/w. They may be standardized in terms of interopacity units or, where appropriate, by
numbers of living or killed bacteria determined by direct cell count or by viable count.

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Bacterial toxoids:
Bacterial toxoids are toxins or material derived therefrom, the toxicity of which has
been reduced to a very low level or completely eliminated by chemical or physical means without
destroying their immunizing potency. The toxins are obtained from selected strains of specific
micro-organisms, grown in media free from ingredients known to cause toxic, allergic or other
undesirable immunological reactions in humans. Toxoids produced by the action of formaldehyde
are known as formol toxoids
Bacterial toxoids may be liquid or may be prepared by adsorbing on
mineral carriers such as aluminium phosphate, aluminium hydroxide or any other suitable
adsorbent; the adsorbed product may be separated, washed and suspended in a saline or other
appropriate solution isotonic with blood.
Bacterial toxoids are clear or slightly opalescent liquids,
colourless or slightly yellow. Adsorbed toxoids may be white or greyish white suspensions or pale-
yellow liquids with a sediment at the bottom of the container. Freeze-dried preparations are greyish
white or yellowish white powders or pellets.
Viral and rickettsial vaccines:
Viral and rickettsial vaccines are suspensions of viruses or rickettsiae and are
prepared from infected tissues or blood obtained from artificially infected animals, from cultures
in fertile eggs, or from cell or tissue culture. Viral vaccines may be live or killed and they may be
freeze-dried. Live vaccines are usually prepared using attenuated strains of the specific organisms.
Killed vaccines may be inactivated by suitable chemical or physical means.
Mixed Vaccines:
Mixed vaccines are mixtures of two or more vaccines. A suitable antibacterial
substance may be added to inactivated or live viral and rickettsial vaccines provided that it has no
action against the specific organisms.

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Impact of genetic engineering on vaccine technology:
The advent of recombinant DNA technology has rendered possible the large-scale production of
polypeptides normally present on the surface of virtually any pathogen. These polypeptides, when
purified from the producer organism (e.g. E. coli, Saccharomyces cerevisiae) can then be used as
‘subunit’ vaccines. This method of vaccine production exhibits several advantages over
conventional vaccine production methodologies. These include:
 Production of a clinically safe product; the pathogen-derived polypeptide now being
expressed in a non-pathogenic recombinant host. This all but precludes the possibility that
the final product could harbor undetected pathogen.
 Production of subunit vaccine in an unlimited supply. Previously, production of some
vaccines was limited by supply of raw material (e.g. hepatitis B surface antigen).
 Consistent production of a defined product that would thus be less likely to cause
unexpected side effects.
A number of such recombinant (subunit) vaccines have now been
approved for general medical use. The first such product was that of hepatitis B surface antigen
(rHBsAg), which gained marketing approval from the FDA in 1986.
Quality control:
Vaccines comply with the following requirements:
(i) Phenol (If present)- Not more than 0.25 per cent w/v.
(ii) Thiomersal (If present)- Between 0.005 per cent w/v and 0.02 per cent w/v.
(iii) Free formaldehyde (If present)- Maximum 0.02 g/l.
(iv) Aluminium (If present)- Not more than 1.25 mg per dose.
(v) Sterility- All vaccines comply with tests for sterility, except that for living bacterial
vaccines, growth of the organism from which the vaccine was prepared is permitted.
(vi)Abnormal Toxicity- All vaccines comply with the test for abnormal toxicity. One human
dose but not more than 1.0 ml is injected intra-peritoneally into each of five healthy mice,

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weighing 17 g to 22 g, and one human dose but not more than 5.0 ml into each of two
healthy guinea pigs weighing 250 g to 350 g.
The preparation passes the test if none of the animals
dies or shows signs of ill-health in seven days following the injection. If more than one
animal dies, the preparation fails the test. If one of the animals die or show signs of ill
health, the test is repeated. The preparation passes the test if none of the animals in the
second group dies or show signs of ill health in the time interval specified.
Storage:
Liquid vaccines are stored at a temperature between 2°C and 8°C and not allowed to freeze unless
otherwise required for some specific vaccines. Freeze-dried preparations must be stored at
temperatures below –20°C. At higher temperatures vaccines deteriorate rapidly.
Some common vaccines:
Bacterial vaccines- Cholera vaccine, whooping cough vaccine, Bacillus Calmette-Guérin (BCG)
vaccine.
Viral vaccines- Oral poliovirus vaccine (OPV; Sabin vaccine), Inactivated poliomyelitis vaccine
(IPV, Salk vaccine), Rabies vaccine, Hepatitis vaccine.
Toxoids: Tetanus toxoid, Diphtheria toxoid adsorbed.
Mixed antigens: Double antigen (DTDA), Triple antigen (DPT).

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References:
 Bhatia, K.N. & Tyagi, M.P. (2011-12). Trueman’s Elementary Biology. New Delhi:
Trueman Book Company. U3-36.
 Walsh, G. (2007). Pharmaceutical Biotechnology Concepts and Applications. West
Sussex, England: John Wiley & Sons Ltd. 400.
 Tripathi, K.D. (2008). Essentials of Medical Pharmacology Sixth Edition. New Delhi:
Jaypee Brothers Medical Publishers (P) Ltd. 879: 881-882:884-885.
 Government of India, Ministry of Health. (2007). Indian Pharmacopoeia 2007 . Delhi:
Manager of Publications. Vol-I:Vol-III.