Antibody against fusarochromanone (TDP‐1) was obtained from rabbits after immunization with TDP‐1 conjugated to bovine serum albumin (BSA). An indirect enzyme‐linked immunosorbent assay (ELISA) using TDP‐1‐ovalbumin conjugate as the antigen coated on to the microtiter plate was used for monitoring the antibody liter. For toxin detection, a direct competitive ELISA in which TDP‐1 was conjugated to horseradish peroxidase (HRP) was used. Competitive direct ELISA revealed that the antibody had about 5.6 and 4.5 times greater binding efficiency for monoacetyl fusarochromanone (TDP‐2) and diacetylated TDP‐1 than TDP‐1. The concentration causing 50% inhibition of binding of TDP‐1‐HRP to the antibody by TDP‐1, TDP‐2 and diacetyl‐TDP‐1 were 2.8, 0.5 and 0.62 ng/ml, respectively. For the analysis of fusarochromanones in wheat and barley, the toxins were first extracted from the commodities with 100% methanol. A small aliquot of the extract was dried, acetylated, diluted in buffer and then analyzed directly by ELISA. The overall recovery for fusarochromanone in the wheat and barley samples spiked with TDP‐1 in the concentration range of 20 to 500 ppb were found to be 97% and 103.4% with cv of 15% and 11.2% for barley and wheat, respectively.