Technical Abstract: There is great incentive to discover biologically active natural products from higher plants that are more effective than synthetic agrochemicals and are environmentally safe. Research emphasis at the US Department of Agriculture has therefore been on the development of alternative approaches to utilizing natural plant products in pest management. Discovery and evaluation of natural product fungicides is largely dependent upon the availability of miniaturized antifungal bioassays. We report on the development of a miniaturized 24-well leaf disc assay for evaluating plant extracts and pure compounds. Compounds applied directly to the leaf surface can be evaluated in a dose-response for fungicidal activity and phytotoxicity. The assay is sensitive to microgram quantities, can determine chemical sensitivity between fungal isolates, and adaptable to complex mixtures, lipophilic extracts, and non-polar compounds. The use of digital imaging and analytical software provided quantitative data and the ability to fine tune the data analysis. Identification of new potential lead compounds can be repeated quickly and on-the-leaf-surface activity can be evaluated in high throughput formats and published in a reasonable time.