Many methods for AFM1 detection exist, but most are time consuming, employ expensive equipment and require experienced personnel. To overcome these problems a membrane-based flow-through enzyme immunoassay has been developed (patent pending). The assay comprised a nylon Immunodyne ABC membrane spotted with anti-mouse antibodies, a plastic snap-fit device, absorbent cotton wool, mouse anti-AFM1 monoclonal antibodies (Mab), and AFB1-horseradish peroxidase (HRP) conjugate. This assay was coupled to an immunoaffinity column (IAC). The visual detection limit was 0.05 ng/g AFM1 in milk. Assay time for IAC clean-up was 12 min, and that for the flow-through assay was 18 min, hence the total assay time was 30 min. This method allows for a rapid screening of milk consignments which do not conform to the maximum permissible limits of 0.05 ng/g AFM1, hence enabling the rejection of such at the farm level. Laboratory validation was done using certified reference materials (CRM) with AFM1 concentrations of < 0.05, 0.09 and 0.76 ng/g. Precision of the assay was high as shown by the high repeatability of the assay results. There were no significant differences in recoveries between standard in buffer and CRM (P > 0.05), and assay responses for these two were highly correlated (99.63%).