Monoclonal antibodies, which are widely applied to the diagnosis and therapy of cancers as well as autoimmune diseases, have been made significant progresses. Currently, the monoclonal antibodies for clinical applications are mostly based on the modified immunoglobulins. However, the variable lymphocyte receptors (VLRs), which are recently discovered as specific antigen receptors of jawless vertebrates, provide a new perspective for the development of antibody reagents and drugs. Compared with immunoglobulins, the antigen-binding specificity, affinity and stability of VLRs-based antibodies are better. In addition, the production technology of antigen-specific monoclonal VLRs has become more and more mature. Therefore, the VLRs have a higher clinical value and may become the next-generation antibody drug. This review focuses on the basic feature, production method and application prospect of VLRs, which provides valuable clues for the applications of VLRs in the field of clinical diagnosis and therapy.