A sensitive sandwich enzyme immunoassay method for measurement of rat α-fetoprotein(AFP) was established by use of affinity purified antibodies to rat AFP. The assay system consisted of filter paper discs with immobilized antibodies and the same antibodies labeled with horseradish peroxidase(POD). The optimal conditions for the assay and for the preparation of POD labeled antibodies were determined. The assay was as sensitive as radioimmunoassay(RIA) and rat AFP at concentrations of 5 to 1280ng/ml was measurable. AFP values obtained by the present method were in good agreement with those by RIA and the correlation coefficient was 0.99.