The Water Framework Directive (WFD), requires European Member States to assess the “ecological status” of surface waters. As part of this, many European countries have developed an ecological quality classification scheme for chlorophyll concentrations as a measure of phytoplankton abundance. The assessment of ecological quality must be based on the degree of divergence of a water body from an appropriate baseline, or ‘reference condition’. It is, therefore, necessary to determine chlorophyll reference conditions for all European lake types. This involves examining how chlorophyll concentrations vary by lake type, in the absence of any nutrient pressures from agriculture or wastewater. For this purpose, a dataset of 540 European lakes considered to be in a relatively undisturbed reference condition has been assembled, including data on chlorophyll concentration, altitude, mean depth, alkalinity, humic content, surface area and geographical region. Chlorophyll was found to vary with lake type and geographical region, and to be naturally highest in low-altitude, very shallow, high alkalinity and humic lake types and naturally lowest in clear, deep, low alkalinity lakes. The results suggest that light and mineral availability are important drivers of chlorophyll concentrations in undisturbed lakes. Descriptive statistics (median and percentiles) of chlorophyll concentrations were calculated from populations of lakes in this reference lake dataset and used to derive lake-type specific reference chlorophyll concentrations. These reference conditions can be applied, through a comparison with observed chlorophyll concentrations at a site, in the assessments of ecological status and provide a consistent baseline to adopt for European countries.