Enhanced infection and replication of porcine reproductive and respiratory syndrome (PRRS) virus in the presence of specific antibody has been demonstrated in vitro and in vivo, a phenomenon known as antibody-dependent enhancement (ADE). ADE is considered to be a significant obstacle to developing effective vaccines for many viruses for which ADE has been reported, since virus-specific antibodies of maternal origin or those conferred by vaccination can facilitate the entry of the virus into target cells, sometimes resulting in increased severity of the disease. In this study, the role of specific PRRS viral epitopes in ADE and/or virus neutralization (VN) was assessed in vitro using 14 monoclonal antibodies (mAbs) to 4 PRRS viral proteins: nucleocapsid (N), matrix (M), glycoprotein (GP) 5, and GP3. Each mAb recognized a distinct epitope on one of these proteins. One-way ADE and VN assays were performed in vitro using homologous PRRS virus isolates in the presence or absence of each mAb. ADE activity was determined by detecting a significant increase of progeny virus yield in porcine alveolar macrophage cultures in the presence of individual mAbs. Neutralizing activity was determined by detecting a significant reduction or complete blocking of virus replication in MARC-145 cells in the presence of individual mAbs. mAbs could be categorized into 3 groups: enhancing, neutralizing and neither. Viral epitopes which are capable of inducing neutralizing antibodies appeared to reside on the M, GP3 and GP5 proteins, while epitopes that may induce ADE-mediating antibody were associated with the N and GP5 proteins. Identification of the viral proteins and antigens and epitopes responsible for ADE- and VN-mediating antibodies may provide the basis for developing efficacious second-generation vaccines for the control of PRRS virus; yet, further epitope mapping remains to be done.