604. Comparative Analysis of Different Methods for Quantitative Evaluation of Neutralizing Antibody to Ebola Virus in a Biosafety Level 2 or 4 Environment

Top of pageAbstract Ebola hemorrhagic fever virus can be modeled in different animal species such as mice, Guinea pigs and nonhuman primates. Several vaccine strategies were evaluated using DNA-, protein-based and/or viral vectors and found to protect mice against challenge with a lethal dose of mouse-adapted Ebola virus. However, only a few strategies involving DNA, Adenovirus or Vesicular Stomatitis virus (VSV), alone or in combination, succeeded at protecting nonhuman primates from Ebola virus aggressive replication. Ebola virus offers thus a unique disease model for the evaluation and testing of different vaccine platforms and/or strategies that can be screened in vivo in conditions of low to high stringency. One major limitation of using Ebola virus as a disease model is the requirement to manipulate the virus in a biosafety level 4 environment (BSL-4) which makes in vitro and in vivo testing less accessible and more time consuming. We compared four different methodologies to evaluate the levels of neutralizing antibody (NAB) from sera of nonhuman primates vaccinated with VSV- or Adenovirus-based vaccine expressing the Ebola envelope glycoprotein (EboGP). The first strategy evaluates neutralization of wild-type Ebola virus by immunoplaques; the second evaluates neutralization of Ebola expressing EGFP inserted as an additional gene in the viral genome; the third evaluates inhibition of plaque formation by an EboGP-pseudotyped VSV virus and the fourth evaluates neutralization of an EboGP-pseudotyped HIV vector. The first two strategies are using a replication competent Ebola virus and were thus performed in a BSL4 laboratory while the other two, using a replication competent VSV or replication defective HIV vector, were done in an enhanced BSL2 environment. Results indicate that the two methodologies using replication competent Ebola virus are slightly more sensitive than with pseudotyped VSV or HIV vector. Ebola expressing EGFP was found the method of choice in the BSL4 laboratory since it can be performed in 48 hours (instead of 6 days for immunoplaque assay) and does not depend on the sensitivity of an antibody/antigen reaction. Overall, each protocol offers advantages and inconveniences that will be discussed.