Protocol for designing a peptide-based multi-epitope vaccine targeting monkeypox using reverse vaccine technology.

Reverse vaccine technology, supported by advancements in immunoinformatics, facilitates the development of multi-epitope vaccines for rapidly evolving pathogens, thereby strengthening the immune defense. Here, we present a protocol for a peptide-based multi-epitope vaccine targeting monkeypox virus (MPXV) using an open-source approach. We describe steps for evaluating physicochemical properties and allergenicity. We then detail procedures for validating pattern recognition receptor (PRR)-binding affinity and stable major histocompatibility complex (MHC) I/II presentation. Molecular dynamics (MD) simulations confirm immune receptor interactions, enhancing vaccine stability. For complete details on the use and execution of this protocol, please refer to Kaur et al.1.