Glacial-relict species of the genus Mysis (opossum shrimps) inhabiting both fresh-water lakes and brackish sea waters in northern Europe show a consistent lake/sea dichotomy in eye spectral sensitivity. The absorbance peak (λmax) recorded by microspectrophotometry in isolated rhabdoms is invariably 20-30 nm red-shifted in "lake" compared with "sea" populations. The dichotomy holds across species, major opsin lineages and light environments. Chromophore exchange from A1 to A2 (retinal → 3,4-didehydroretinal) is a well-known mechanism for red-shifting visual pigments depending on environmental conditions or stages of life history, present not only in fishes and amphibians, but in some crustaceans as well. We tested the hypothesis that the lake/sea dichotomy in Mysis is due to the use of different chromophores, focussing on two populations of M. relicta from, respectively, a Finnish lake and the Baltic Sea. They are genetically very similar, having been separated for less than 10 kyr, and their rhabdoms show a typical lake/sea difference in λmax (554 nm vs. 529 nm). Gene sequencing has revealed no differences translating into amino acid substitutions in the transmembrane parts of their opsins. We determined the chromophore identity (A1 or A2) in the eyes of these two populations by HPLC, using as standards pure chromophores A1 and A2 as well as extracts from bovine (A1) and goldfish (A2) retinas. We found that the visual-pigment chromophore in both populations is A1 exclusively. Thus the spectral difference between these two populations of M. relicta is not due to the use of different chromophores. We argue that this conclusion is likely to hold for all populations of M. relicta as well as its European sibling species.