Abstract: A reproducible enzyme-linked immunosorbent assay(ELISA)using Marek’s disease virus(MDV)-infected cells for the detection of antibodies to MDV is described.The optimum number of MDV-infected chicken embryo fibroblasts(CEF)was 5 × 104/well,and test sera were positive at 1:400 dilutions.Compared with a purified virus preparation, MDV-infected CEF produced high specific and low nonspecific reactivities. wells coated with whole cells could be stored at 4 ℃ or-20℃ for at least 3 months without loss of reactivity, with antibody-negative sera, the cutoff absorbancy was 0.20 units.The ELISA was 20-to-40-fold more sensitive than indirect immunofluorescence.Homologous combinations of antisera in wells coated with CEF infected with different MDV serotypes were more reactive at higher dilutions than that were heterologous combinal ions. The procedure described is specific and suitable for large-scale screening of both chicken and monoclonal antibodies against MDV.