The specific interaction between antigen and antibody is determined by the specific complementary recognition between antigenic determinant and epitope motif on the surfaces of antibody and antigen, respectively. B cell epitope mapping involves both the fine location of B cell epitopes (location of the specific motifs on an antigen recognized and bound by antibodies) and the depiction of the distribution of all or nearly all epitopes of an antigen on its primary or secondary structure. Mapping of B cell epitopes constitutes a primary basis for the development of epitope-based vaccines and therapeutic epitope specific antibodies and the establishment of immunological diagnostic methods. So far, a number of experimental methods have been developed to map the B cell epitopes recognized by antibodies or to depict the map of epitopes on an antigen. B cell epitope mapping methods based on X-ray crystallography of antigen-mAb (monoclonal antibody) complex and screening of the mutant libraries of an antigenic protein or its fragments can reveal the key residues of antigen specifically bound by mAb at amino acid or even atom level. Other epitope mapping methods, such as peptide library screening technology using ELISA, are seldom used to identify the minimum epitope motifs because only the antigenic fragments can be obtained by these methods. On the other hand, the improved biosynthetic peptide method is usually used for the minimal motif identification and fine mapping of B cell epitopes. It is quite common that more than one methods were employed to fine map B cell epitopes due to the respective limitations of each method. This review discusses and compares different experimental methods commonly used in B cell epitope mapping and aims to assist researchers to design the most suitable protocol to map their B cell epitopes. The applications of B cell epitope mapping in animal disease prevention and control are also reviewed.