Worldwide concerns about fossil fuel depletion and energy security have recently triggered a research interest in biodiesel, which is renewable, biodegradable, and has several other advantages as an alternative to petro diesel. However, biodiesel may cause engine problems, especially fuel filter plugging, associated with its use in cold weather conditions. Trace contaminants such as glycerin, saturated monoglycerides (SMG), and soap compromise cold weather performance of biodiesel. A cold soak filtration test was recently included in the U.S. specifications for biodiesel (ASTM D 6751-09) to evaluate biodiesel cold weather performance. Canola seed has good potential to be a locally important biodiesel feedstock because of its high yield (1500 to 2200 kg/ha) and oil content (40 to 50%, Brassica napus L.), as well as a suitable fatty acid profile for good cold weather performance. For a plant breeding program evaluating canola biodiesel quality traits, rapid preparation of biodiesel samples and assessment of its quality is important. In this work, an in situ alkaline transesterification method was adopted for preparing canola biodiesel. It was found that the biodiesel yield via this method was improved by reducing seed moisture from 6.7% to 0% after oven-drying. The resulting biodiesel had qualities comparable to or better than biodiesel prepared through the conventional alkaline transesterification. Only a limited amount of seed from new canola lines is typically available in a plant breeding program; obtaining the required volume of biodiesel for evaluating cold soak filterability (300 mL) is not possible. In order to rapidly screen canola breeding lines for B 100 quality, cold soak filterability must be assessed with reduced volumes of biodiesel. Therefore, this study evaluated the impact of SMG, glycerin, and soap on cold soak filterability. Biodiesel filtration time rapidly increased to unacceptable levels and became much less reproducible when the SMG concentration was raised above 0.28%. A regression model was generated to predict the filterability of biodiesel against the concentrations of trace contaminants. A downscaled model of the filtration test with a reduced volume of biodiesel sample (25 mL) was also tested and calibrated. The in situ transesterification method saved 30% operator time compared with the conventional method. By combining the downscaled cold soak filtration test, the goal of analyzing 40 biodiesel samples/wk was achieved.