ALTEX :: Alternatives to Animal Experiments

1998, Supplement

How to Prove Complete Virus Inactivation in Rabies Vaccines: a Comparison of an In Vivo to an In Vitro Method

Stephanie A. E. Blum, Manuela Braunschweiger, Beate Kraemer, Petra Ruebmann, Karin Duchow, and Klaus Cussler

Paul-Ehrlich-Institut, D-Langen

SUMMARY

At present, the complete inactivation of rabies virus in rabies vaccines ad us. vet. is proven by an animal experiment which causes severe suffering, the intracerebral injection of mice. This animal experiment is not validated yet. We have quantified the sensitivity of the mouse test and examined whether the animal experiment may be replaced by the immunofluorescence assay (IFT) as an in vitro method. Detection limits of both assays were determined depending on the examined product, i.e. prior to and after the addition of adjuvants and preservative, respectively. Furthermore, symptoms of the rabies disease were recorded and their severity was classified on a range of 1-5. Symptoms of rabies-infected mice were clear and highly specific. Symptoms classified as ≥ 2 in context with a loss of ≥ 15% of the initial weight were defined as humane endpoints of the disease. The quantitative detection of active virus was not inhibited in the presence of even high concentrations of inactivated virus. The detection limit of the mouse test was 10 viruses ml-1 independent of the examined product. The detection limit of the IFT prior to the addition of adjuvants and preservative was 10 viruses ml-1 as well. After the addition of these substances, the detection limit rose to 103 viruses ml-1. Advantages and disadvantages of the mouse test and IFT are discussed.

Keywords: rabies virus, virus inactivation, safety test, immunofluorescence assay, intracerebral injection