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YELLOW FEVER VACCINE, LIVE

Category Active immunizing agent.

      Live Yellow Fever Vaccine is a freeze-dried preparation of live attenuated 17D strain of yellow fever virus grown in embryonated chicken eggs.

      The vaccine, reconstituted as stated on the label, complies with the requirements stated under Vaccines, with the following modifications.

Description Slightly dull, light orange-coloured, flaky or crustlike, desiccated mass. When reconstituted, it is clear or slightly opalescent and light orange liquid.

Strength available Not less than 1 × 103 mouse LD50 or its equivalent in PFU per 0.5 ml.

Dose Subcutaneous, 0.5 ml.

Contra-indication

1. It is contra-indicated in individuals with a history of anaphylactic hypersensitivity to eggs or chicken protein.

2. It is contra-indicated in infant younger than 4 months of age.

Warning

1. It is not recommended in infants under 6 months of age except in high-risk areas.

2. Risk-benefit should be considered if it is to be administered during pregnancy.

3. Risk-benefit should be considered if it is to be administered in individuals with history of hypersensitivity to eggs or chicken protein.

Expiration date When stored under the prescribed conditions, the expiration date is not later than 2 years after the date of the last satisfactory test for virus titre.

Packaging and storage Yellow Fever Vaccine should preferably be stored at all time at the temperature below 5º, protected from light. Do not freeze.

      When reconstituted, the vaccine should be used immediately.

Labelling Complies with the “General Information for Biological Products”, p. 177. In addition the label on the container states (1) the strain of virus used in preparation of the vaccine; (2) that the vaccine has been prepared in chick embryos; (3) the minimum virus concentration; (4) that contact with disinfectants is to be avoided; (5) the period of time within which the vaccine is to be used after reconstitution.

Identification When the vaccine is mixed with an appropriate amount of specific yellow fever antiserum, there is a significant reduction in its ability to infect susceptible cell cultures.

Ovalbumin Not more than 5 μg of ovalbumin per human dose, determined by a suitable immunochemical method (Appendix 14.5).

Bacterial endotoxins When tested as described in the “Test for Bacterial Endotoxins” (Appendix 8.5), it contains less than 5 Endotoxin Units per human dose. 

Thermal stability Maintain not less than three vials of the vaccine in the dry state at 37º for 14 days. Determine the virus concentration as described under Assay in parallel for the heated vaccine and for unheated vaccine. The difference in the virus concentration between unheated and heated vaccine does not exceed 1.0 log and the virus concentration of the heated vaccine is not less than the number of PFU equivalent to 1 × 103 mouse LD50 per human dose.

Assay

      Titrate for infective virus in cell cultures. Use an appropriate virus reference preparation to validate each assay.

      The virus concentration is not less than the equivalent in PFU of 1 × 103 mouse LD50 per human dose. The relationship between mouse LD50 and PFU is established by each laboratory and approved by the competent authority.

      Before assay, the reconstituted vaccine shall stand at a temperature between 20º and 30º for 20 minutes. Appropriate serial dilutions of the reconstituted vaccine are made in diluent for yellow fever virus which has been demonstrated to be free of yellow fever virus inhibitors.

      The method shown below, or another suitable technique, may be used to determine the mouse LD50 and the PFU.

     MOUSE LD50 TECHNIQUE

      A strain of mice highly susceptible to yellow fever virus, aged 4 to 6 weeks, are injected intracerebrally with 0.03 ml of vaccine dilution. Groups of at least six mice are used for each dilution. The series of dilutions to be used should result in mortality rates which span from the range of 0 to 100 per cent. Inoculation of the mice should be performed immediately after the dilutions have been made. All deaths are recorded during a period of 21-day observations. Mice dying from unrelated causes are removed from both the numerator and denominator of mortality calculations. Mice paralyzed on day 21 are counted as alive.

      The mouse LD50 is the quantity of virus suspension estimated to produce fatal, specific encephalitis in 50 per cent of intracerebrally inoculated mice.

      CELL-CULTURE TECHNIQUE

      Use either Method I or II.

      Method I Monolayers of Vero or other suitable cells are prepared in multi-well plates. At least three wells are inoculated with a virus dilution. After incubation for 1 hours at 37º±1º, the virus dilution is replaced by a suitable overlay medium consisting of Leibovitz medium No. 151 or Minimum Essential Medium (MEM)1 , 5 per cent of fetal bovine serum, and 1.6 per cent of carboxymethylcellulose (low viscosity sodium salt) or other suitable medium. The plates are incubated in a 5±1 per cent carbon dioxide incubator at 37º±1º for 5 days. On the sixth day, the plates are drained, washed 

with saline TS, stained with a 1 per cent w/v solution of naphthalene black or any other suitable stains and thoroughly rinsed with tap water. The plaques are then counted and virus titre is calculated.

      Method I Equal amounts (0.2 ml) of a Vero-cell suspension (approximately 6 × 105 cells/ml) and virus dilution in Leibovitz medium No. 15 or MEM or other suitable medium are placed in each of the 16-mm flatbottomed wells in sterile trays suitable for cell culture. The trays are sealed and incubated for 4 hours at 37º±1º in a 5±1 per cent carbon dioxide incubator. After incubation, 0.4 ml of overlay medium, consisting of Leibovitz medium No. 15 or MEM medium, 3 per cent of fetal bovine serum, and 1.6 per cent of carboxymethylcellulose (low viscosity sodium salt), is added to each well. The trays are resealed and incubated at 37º±1º in a 5±1 per cent carbon dioxide incubator for 5 days. On the sixth day, the trays are drained, washed with saline TS, stained with a 1 per cent w/v solution of naphthalene black or any other suitable stains and thoroughly rinsed with tap water. The plaques are then counted and virus titre is calculated.


1Use a commercially available medium.

MONOGRAPHS • YELLOW FEVER VACCINE, LIVE
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หมายเหตุ / Note : TP II 2011 PAGE 281-283