A method is described for the analysis of grain samples for the presence and quantitative estimation of sterigmatocystin. Initial extraction with acetonitrile-water (9 + 1) containing KCl is followed by partition of the extract first against hexane and then against CHCl3, and, if needed, silica gel column chromatography. This sequence provides an extract in which the sterigmatocystin can be estimated by fluorescence intensity comparison on TLC plates against a pure sterigmatocystin standard. Visualization of the sterigmatocystin on the TLC plates is enhanced by an AlCl3 spray reagent. After preparative TLC, the identity of sterigmatocystin is confirmed by formation of an acetate derivative and of a derivative formed by treating the suspected extract with aqueous acid.