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Rift Valley fever virus is a mosquito-borne virus which is associated with acute hemorrhagic fever leading to large outbreaks among ruminants and humans in Africa and the Arabian Peninsula. RVFV circulates between mosquitoes, ruminants, camels and humans, which requires divergent amplification and maintenance strategies that have not been fully explored on the cellular and molecular level. We therefore assessed monoclonal antibodies for their applicability to monitor the expression pattern and kinetics of viral proteins in different RVFV infected cell species. Sequences of RVFV vaccine strain MP-12 were used in a bacterial expression system to produce recombinant non-structural proteins directed to NSs and NSm. After immunization of balb/c mice a set of monoclonal antibodies were generated and extensively characterized. The kinetics of RVFV proteins in vertebrate (Vero76) and mosquito-derived (C6/36) cells were evaluated with monoclonal antibodies against the nucleocapsid protein (NP) and the glycoproteins (Gn and Gc) as well as with the newly generated NSs and NSm derived monoclonal antibodies. Significant differences of viral protein distribution and accumulation in vertebrate compared to mosquito-derived cells could be demonstrated. Differences were observed for the nonstructural NSm and most intriguingly for the NSs protein indicating significant divergency of replication strategies of RVFV in Vero 76 cells and C6/36 cells. The described monoclonal antibodies are therefore powerful tools to elucidate the discrepancies of virus replication and interaction within the mammalian host compared to the mosquito vector. [ABSTRACT FROM AUTHOR]