Previous studies have investigated the impacts of biofuel usage on the performance, drivability and durability of modern diesel engines and exhaust after-treatment systems including test work with different types, concentrations and mixtures of bio fuel components. During this earlier work vehicle fuel filter blocking issues were encountered during a field trial using various types of EN 14214 compliant Fatty Acid Methyl Ester (FAME) blended into EN 590 diesel. This paper summarises a subsequent literature review that was carried out looking into potential causes of this filter blocking and further work that was then carried out to expand on the findings. From this, a laboratory study was carried out to assess the increase in fuel filter blocking tendency (FBT) when various FAMEs from mixed sources were blended into EN 590 diesel at different concentrations, including levels above those currently allowed in the European market. Results have indicated that there are certain ratios of FAME from different sources which interact antagonistically to form higher levels of filter deposits. Using this knowledge, experimental fuel formulations for a three-vehicle field trial were selected to assess engine and exhaust after-treatment system durability issues seen when blends made from mixed FAME sources are used. All three vehicles encountered fuel filter blocking issues during relatively early stages of the trial. Deposits from the blocked filters have been analysed which has confirmed steryl glucosides, amongst other species, to be a strong contributory factor, which is consistent with the findings of the literature review and a trace component laboratory investigation also carried out as part of this study. This paper finds that increased filter blocking tendency of mixed FAME blends is likely due to the complex interaction of trace components found in the various FAME types and is also strongly affected by storage temperatures, time in storage and properties of the base diesel.