A micro-ELISA technique was developed for the detection of Epstein-Barr virus (EBV)-determined antigens. The enzyme-linked immunosorbent assay (ELISA) was applied with peroxidase-protein A to detect the antigens adsorbed to micro-ELISA plates. Human and rabbit antisera containing antibodies to known EBV components were used as reagents. The early antigen (EA) complex, associated with the viral cycle, was readily detected in extracts of n-butyrate- or n-butyrate + TPA-induced cells. The nuclear antigen, EBNA, could be unequivocally detected only after the partial purification of the antigen by DNA cellulose chromatography. EA (and VCA) could be separated by chromatofocusing of induced cell extracts into several fractions detected by the micro-ELISA technique. This indicates that the purification of individual antigens of the EA complex can be monitored by ELISA.