Stage #1: input data gathering and preparationProteome retrievalsequences ideally fromall strains or species of target organismCreate core proteomeRemove proteins that are...paralogous or duplicatedhomologous to vaccine recipient sequences toxicallergenicprotein sequences of core proteome-Stage #2: predicting proteins exposed to immune system Classical reverse vaccinologyPredict informative protein characteristics...[Antigenicity] [Subcellular localisation] [Transmembrane domains] [Function] [Signal peptides] [Virulence] [Adhesion] [Physico-chemical properties]sequences of antigenic proteins1-Stage #3: predicting epitopes (immunoinformatics)Epitope selection based on...1 Helper T-lymphocyte (HTL) Cytotoxic T lymphocytes (CTL) B-cell epitopesbinding affinity epitope density population coverageVaccine constructStage #4: computationally verify vaccine construct- Check for... Antigenicity, Allergenicity, Toxicity, Solubility, StabilityProtein-protein interactionCodon optimization In silico cloning3D structure modelling Structure refinementMolecular docking with receptor Molecular dynamics simulation | Binding free energy calculationsimmune system simulationMulti-epitope Vaccine for laboratory testing