Verification of the rabies virus glycoprotein lower limit of immunogenicity by serological assay.

Rabies lethality is close to 100% and annually 15 million people receive post-exposure prophylaxis. Testing for vaccines against this zoonosis should ensure its quality. A standardized test by the National Institutes of Health (NIH) test, based on mice immunization and challenge, has been used to determine the potency of vaccine lots. It has several disadvantages, the main one being its significant variability. Several in vitro methods like an Enzyme-linked immunosorbent assay (ELISA) have been proposed based on the quality and quantity of glycoprotein (Glptn) of rabies virus, but may also present limitations such as low sensitivity, instability and imprecision. The estimate of immunogenicity based on neutralizing antibody titer (Nab) evaluated by a serological test (ST) such as the Modified Rapid Fluorescent Focus Inhibition Test (mRFFIT), is not yet efectively applied for human vaccine. Nevertheless, a Nab concentration can be used as a predictor of clinical efficacy of this product in vaccinated humans, so, that can be applied in estimating the vaccine potency. The aim of this study was to verify the lower limit of immunogenicity of the viral Glptn content in mice using mRFFIT. The lower Glptn content by ELISA able to induce Nab response was determined. The results were correlated and demonstrated that ST was able to determine the Glptn immunogenicity lower limit. Our findings suggest that a test based on rabies Nabs may represent an additional alternative for the evaluation of rabies vaccines.

Development and pre-validation of a quantitative multi-dose serological assay for potency testing of inactivated rabies vaccines for human use.

It is mandatory to ensure the quality of biological products used in the prevention of rabies, a zoonosis with nearly 100% lethality. Fifteen million people receive post-exposure prophylaxis yearly. The vaccine batches are assessed by the National Institutes of Health (NIH) test which has several disadvantages such as significant variability and animal welfare issues. The estimation of immunogenicity based on titration of neutralizing antibodies (NA) is not applied to the human vaccine yet. Despite this, a satisfactory concentration of NA (0.5 IU/ml) can be used as a predictor of the clinical efficacy and for estimating rabies vaccine potency. The objective of this study was to develop and pre-validate a Serological Potency Test (SPT) using the modified Rapid Fluorescent Focus Inhibition Test (mRFFIT) to determine the potency of rabies vaccines for human use, demonstrating its relevance and reliability. The results show good agreement between the potencies determined by the SPT and the NIH test. The assay was able to distinguish between potent and sub-potent lots of vaccines. The results demonstrated that SPT is a viable candidate for validation and inclusion in pharmacopeias as a reduction and refinement for the NIH test.