Real-time fluorescence monitoring has been mostly performed in marine systems, with little progress being made in the application of fluorescence excitation emission matrix (EEM) spectroscopy, especially for freshwater monitoring. This paper presents a two weeks experiment where real-time fluorescence EEM data have been obtained for Bourn Brook, Birmingham, UK, using an in-situ fibre-optic probe. Fluorescence EEMs were measured every 3 min for two weeks, with control 'grab' samples every hour analyzed for fluorescence EEMs as well as pH, conductivity and dissolved organic carbon. Comparison of real-time and control samples showed an excellent agreement, with no evidence of fibre-optic probe fouling. EEMs of different character were identified using self-organizing maps, which demonstrated seven clusters of fluorescence EEMs which related to the intensity of fluorescence and relative intensities of peak T-1 and T-2 vs. peak C and peak A fluorescence. Fluorescence intensity of peaks A and C were observed to increase with rainfall, and a diesel pollution event was detected through an increase in T-2 fluorescence. (C) 2010 Elsevier Ltd. All rights reserved.