Two-photon laser scanning microscopy of lotic biofilms (excitation and emission see Table 1). The first data set (a, b) showing a comparison between one-photon (a) and two-photon (b) excitation of autofluorescence from cyanobacteria and algae from the same location. The second data set (c–f) showing a nucleic acid stained biofilm sample. The single channel signals showing (c) SYTO 40 stained bacteria in the green channel, (d) autofluorescence of cyanobacteria in the red channel, (e) chlorophyll signal of cyanobacteria and algae in the far red channel and (f) the resulting overlay of all three channels (green=bacteria stained with SYTO 40, pink=autofluorescence of cyanobacteria, blue=autofluorescence of algae, white=red/green/blue overlay of cyanobacteria).