Abstract
Background: The development of neutralizing antibodies against HIV-1 is of pivotal importance for the development of an AIDS vaccine. We immunized mice with fusion complexes and elicited antibodies with neutralizing activity against heterologous HIV-1 isolates (Zipeto et al., Microb Infect 2006). Methods: We prepared murine hybridomas from mice whose sera showed the highest neutralizing activity. Their supernatants were tested for reactivity against cells expressing CD4 and CCR5, gp120/41, and the gp120/CD4 complex formed by capture ELISA. Hybridoma antibodies with no reactivity against HIV-1 receptors were selected. IgGs were purified and tested for their neutralizing activity (1-5 μg/mL) using both the TZM-bl neutralization assay (Li M. et al, J Virol 2005) and pNL4-3.Luc.R-E- based pseudoviruses on U87R5 cells with the standard group B Env panel. Specificity was tested against the VSV-G envelope protein. The neutralizing activity of selected antibodies was confirmed using the PBMC-based neutralization assay (Polonis VR et al, Virol 2008). Results: We screened 150 different hybridoma groups; 8% showed a neutralizing activity higher than 40% and 1 between 50 and 80% against the different pseudoviruses tested, similar or higher than the Tri-mAb positive control. Cells from this hybridoma group were cloned; 7% showed a neutralizing activity higher than 40% and 2% higher than 50%. Among the latter as many as 44% showed a neutralizing activity higher than 75% against the different pseudoviruses tested. Conclusions: Hybridomas produced from mice immunized with fusion complexes secrete antibodies neutralizing a panel of HIV-1 clade B Envs. We are screening monoclonal antibodies for their broad spectrum neutralizing activity against HIV-1. The isolation of such monoclonal antibodies will be of great interest for the development of an AIDS vaccine.
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More From: JAIDS Journal of Acquired Immune Deficiency Syndromes
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