An indirect, double antibody, microtitration plate ELISA technique was used to assay aflatoxin B, in samples of peanuts, Brazil nuts, almonds, hazelnuts and walnuts. The minimum dilutions of nut extracts required to prevent interferences were of 1 in 10 for almonds; of 1 in 20 for Brazil nuts and peanuts; and of 1 in 50 for hazelnuts and walnuts. The ELISA procedure was validated for aflatoxin B, detection in the shelled nuts referred to above by determining the percentage of this toxin recovered from artificially spiked nut samples. The results confirmed that this technique can be used for aflatoxin B1determination in these nuts. The comparative studies undertaken using naturally contaminated peanut samples also led to the conclusion that this ELISA protocol can be recommended as an alternative to the already adopted thin‐layer chromatography (TLC) method. It was also observed that placing peanut samples under a UV light is not an advisable method of ascertaining aflatoxin B1contamination of these nuts, as levels of 0.6 μg g‐1were not detectable.