Integration of an enzyme-linked immunosorbent assay (ELISA) with conventional chromatographic methods proved the versatility of ELISA as a research tool and allowed for rapid assessment of aflaxtoxin in individual cottonseeds, parts of cottonseed, and composite samples of seeds taken from individual cotton bolls. Aqueous acetone was substituted for methanol in the extraction procedure prescribed by ELISA. The substitution allowed the use of a common extract for all analytical methods. An aliquot of the extract was used to screen samples by ELISA. Negative samples were identified, and toxin levels between 1 and 70 ng/g were quantitated by ELISA. Samples with toxin levels beyond the upper limit of detection by ELISA were then subjected to more time-consuming conventional cleanup prior to quantitation by liquid chromatography (LC) or thin-layer chromatography (TLC). Toxin levels detected by LC or TLC ranged from 100 to 845,000 ng/g sample. The screen by ELISA detected large numbers of toxin-negative cotton bolls or individual seeds in minimum analysis time. The combination of techniques verified the presence of seed with no toxin adjacent to toxin-containing seed in the same lock. Toxin-negative portions of individual seed with high toxin in another portion were identified for the first time. Integration of techniques provided needed information on distribution patterns of aflatoxin in cotton so that preventive measures can be developed.