Preparation of Antiserum to Apple stem pitting virus Using in Silico Prediction of Antigenic Epitopes

A sensitive antiserum is needed for the detection of Apple stem pitting virus (ASPV), one of the most important latent viruses that infect fruit trees. We have studied many properties of coat protein, such as the antigenic index, α‐helix, β‐sheet, β‐turn, coil structure, hydrophilicity, surface probability and flexibility and analysis with several software algorithms. Based on the rules for locating the antigenic epitopes in the regions including β‐turns and coil structures with the high hydrophilicity and surface probability, the predicted epitopes were located in the region of amino acid positions 4–18, 100–114, 400–414, respectively. Two linear synthetic peptides (CRGYEEGSRPNQRVLP and CTGGKIGPKPVLSIRK) were prepared and conjugated with carrier protein. The antisera, designated 1468 and 1469, were obtained by immunizing rabbits. The antibody produced a strong immuno‐reaction with the expression product of the ASPV coat protein gene in Escherichia coli. By testing ten apple samples, ASPV could be detected by Protein A Sandwich ELISA using antiserum 1468, but only some positive samples could be detected with antiserum 1469. To our knowledge, this is the first report of the preparation of antiserum to a pome fruit virus using the antigenic epitopes method.