The test is performed as a direct ELISA to detect Ebola Virus (EBOV) viral matrix protein 40 (VP40) and nucleoprotein (NP). Diluted samples, reference, and controls are incubated in microwells coated with purified EBOV antigen specific polyclonal antibodies. Incubation allows the antigen present in the samples to react with the immobilized antibodies. After the removal of unbound serum or plasma proteins by washing, purified EBOV antigen specific polyclonal antibody conjugated to horseradish peroxidase (HRP) is added to complex with the bound antigen. Following another washing step, the bound anti-EBOV HRP conjugate is assayed by the addition of tetramethylbenzidine (TMB) and hydrogen peroxide (H2O2) chromogenic substrate. The intensity of color development in the wells is proportional to the serum or plasma concentration of EBOV antigen. Results are obtained by reading the OD (optical density or absorbance) of each well in a spectrophotometer. Antigen concentration can be estimated from a serial dilution curve prepared from the Reference provided in the kit. Positive and Normal serum controls are provided. It is recommended that the user establish a cut-off for the study population using at least 100 normal (non-febrile) serum samples.