Sclerotinia sclerotiorum is a major pathogen of many crops, including oilseed rape (Brassica napus), and there is keen interest worldwide to identify Brassica genotypes with resistance to this pathogen. However, field testing to identify resistance in B. napus germplasm is expensive, time-consuming and at times unreliable due to variability in field environmental conditions and plant architecture. To address this, we aimed to examine the feasibility of utilising for B. napus a cotyledon test already developed for Sclerotinia disease on legumes. Initially, cotyledons of 32 B. napus genotypes were drop-inoculated using macerated mycelium (10(4) mycelial fragments/mL) under controlled environmental conditions. Significant differences were recorded between B. napus genotypes, and the experiment was repeated twice using genotypes selected from the first experiment. Certain genotypes responded with a distinct hypersensitive reaction (lesions < 1 mm diameter), either always (cv. Mystic) or frequently (cv. Charlton), which is the first report of this phenomenon in the B. napus-S. sclerotiorum pathosystem. Responses of genotypes between the three screening experiments were significantly and positively correlated. Results obtained in the first experiment were compared with those from our earlier field screening for stem rot that utilised the same strain of S. sclerotiorum and the same B. napus genotypes. In particular, there was a significant positive correlation (r = 0.62, P < 0.01) between published field data for stem rot and our cotyledon test results across genotypes in common. This indicates the usefulness of this cotyledon assay to provide a relatively reliable indication of field performance of genotypes. We believe that this is the first report demonstrating that a cotyledon assay can be successfully applied to rapidly differentiate the reactions of B. napus genotypes against S. sclerotiorum.