Air pollution, in particular ambient particulate matter (PM), can be linked to a variety of different health effects, and a major contributor to the PM pollution is exhaust from diesel engines and other vehicles. In the global drive towards finding sustainable and clean bio-based alternative fuels for the transport sector, biodiesel is one of the most established alternative. However, there is considerable variation in emission data for biodiesel, preferably explained by influences of engine technology and operating conditions as well as dilution sampling strategy. In this study the focus was therefore to study the effects of dilution conditions on the particle formation and size distribution in the exhaust emissions from an off road engine, when introducing RME biodiesel in comparison to standard petro diesel. Particle size distribution and number concentration were measured on-line with the use of a fast mobility spectrometer, during a transient operation and without engine modification. Differences in particle characteristics were elucidated in the raw exhaust versus diluted exhaust at two subsequent sampling points with different dilution ratios. In addition, the influences on the exhaust particle properties of changing the lubrication oil was investigated. It was found that biodiesel in general generated more nucleation mode particles then petro diesel, and after the oil exchange the total particle number concentration was increased even more. It was also seen that the custom-built dilution setup favors generation of nucleation mode particles, which is in line with real life conditions in chase and road side experiments. However, when using heated primary dilution and a heated line in the raw exhaust the formation of nucleation mode particles was suppressed. Overall, it was concluded that the introduction of the biodiesel, and potentially other renewable fuels, can in a considerable way change the exhaust particle emission and characteristics. This could have implications for the assessment of exhaust from engines running on biodiesel fuels, especially when introducing biodiesel in existing and older engines.