Many biological preparation become unstable during storage and thus their safety and efficacy are reduced. Proteins and other macromolecules may be sensitive to heat, light, radiation, and environmental changes, or they may interact with the container materials or other components of the vaccine mixture.

Determining these relationships and optimising stability from the time of production to administration is an important part of vaccine development. The potency of vaccine may reduce as a function of the time elapsed since production even if optimal conditions are maintained.

The stability of vaccines may undergo significant changes after being exposed to temperature fluctuations during handling or storage. Thus, their stability characteristics should be determined through testing, and the storage and handling conditions should be defined to ensure that minimum standards of potency, identity, and purity continue to be met throughout the shelf-life.

Bacillus Calmette-Guerin or BCG Vaccine

Bacillus Calmette-Guerin (BCG) vaccine provides immunisation against tuberculosis. It is prepared using live attenuated (weakened) bovine tuberculosis bacillus strain of Mycobacterium bovis, which has lost its virulence in human.

Preparation

A pure culture of an authentic strain of attenuated Mycobacterium tuberculosis (of Calmette and Guerin) is grown for 2 weeks on a suitable media. The attenuated strains are collected and suspended in a liquid medium designed to preserve the antigenicity and viability of vaccine. It is dispensed either in the freeze dried form or in liquid form.

Method of Administration

Before administering the BCG vaccine, a tuberculin skin test should be done in . the patients (except in neonates). If a patient with a positive tuberculin reaction is administered with the vaccine, he/she is at a high risk of severe local inflammation and scarring. It is a false belief that people showing positive tuberculin reaction do not require BCG vaccine as they already possess immunity against it; however, these patients should further undergo a screening for the presence of active tuberculosis.

BCG is administered as a single injection given via intradermal route at the point of insertion of the deltoid muscle. Accidental subcutaneous administration develops a localised abscess (known as a BCG-oma), which further leads to ulcerations and is treated with antibiotics.

Storage Conditions and Stability

BCG vaccine is freeze-dried and should be stored in dark places at 2−8°C temperature. It is more stable if stored at temperatures as low as −20°C. It should be protected from light. The diluent should be stored in a cool place. The diluent should not be frozen or else the vaccine will denature due to freezing and thawing. During transportation, the vaccine should be kept on ice in a cool bag.

BCG is very sensitive to heat, light, and neon light. Under suitable storage conditions, the effectiveness of the vaccine can be maintained even after two years of manufacture. The recommended maximum time for which BCG vaccine can be stored and consumed (its expiry) is two years from the date when the last potency test performed yielded satisfactory results. However, the vaccine remains effective for 2 years only when stored in dark under the required temperature.

Pertussis Vaccine (Whooping Cough Vaccine)

Pertussis (or whooping cough) is an acute, communicable disease, caused by Bordetella pertussis (a small, non-motile, and gram-negative bacillus). This bacterium takes 7-10 days for incubation; a catarrhal stage follows the peculiar paroxysmal cough that ends in a whooping inspiration.

Pertussis can be prevented if the infants are immunised against it from the time they are of three months. The patients of whooping cough are provided symptomatic and supportive treatment. Pertussis vaccine is a whole cell preparation obtained from the killed Bordetella pertussis. It is also the third constituent of the DTP-triple vaccine.

Preparation

B. pertussis isolated from an infected patient is dried and cultured under specific culture conditions. This phase of collecting a sterile suspension of B. pertussis is the Phase-I.

The cup-plate method is used for isolating the microorganisms from the patient, which are then collected once they are in a smooth and virulent form, generally referred to as Phase-I. It has been observed that microorganisms in Phase-I appear smooth, but when grown on an unfavourable media, a S→R variation takes place with the loss of important antigens. Phase-IV corresponds to a fully developed rough form.

Bordet-Gengou blood agar media is used for growing the microorganisms and then incubating them for 24-74 hours.

Then it is suspended in NaCl-containing injection, and the obtained suspension is centrifuged. The bacterial mass is then shifted to NaCl injection consisting of 0.01−0.02% thimerosal to prevent the growth of bacteria, without altering the antigenic properties of the prepared vaccine.

Standardisation

Each dose of 0.5ml should have 7.5μg of pertussis antigens, which are expressed as protein nitrogen. The obtained vaccine constitutes inactivated PT and FHA proteins each in 23.4μg amounts.

Storage Conditions and Stability

The concentrated preparation obtained is stored for 3 months in a refrigerator to reduce its toxicity. After 3 months, the preparation is diluted with adequate amounts of NaCl injection so that its opacity is greater than twice to that of the standard preparation in a final concentration of 0.01% of thimerosal. Thus, thimerosal kills the present microorganisms. This vaccine does not require heat treatment.