Mycotoxins are naturally occurring toxic chemical compounds produced by fungi infesting agricultural crops both during crop growth and storage. Such secondary metabolites, when ingested, can produce toxic syndromes in humans. This study is the first survey that documents the occurrence of mycotoxins in stored barley in Tibet Autonomous Region [P.R. China]. Twenty-five samples of barley collected from Tibet were analyzed for the presence of aflatoxins, fumonisins, ochratoxins, zearalenone, deoxynivalenol, and T-2 toxin using an easy, sensitive, competitive direct enzyme-linked immunosorbent assay. Ninety-six percent of the samples were contaminated with zearalenone at concentrations ranging from 25 to 270 µg/kg. Seventy-six percent of the samples were contaminated with T-2 toxin at concentrations ranging from 1 to 163 µg/kg. In contrast, deoxynivalenol was observed in only 12% of the samples, with toxin concentrations ranging from 25 to 270 µg/kg. Aflatoxin was observed in only 4% of the contaminated samples.