Vinyl-pyridinium triphenylamines: Novel far-red emitters with high photostability and two-photon absorption properties for staining DNA

A series of mono-, bis- and trisvinyl-pyridinium triphenylamines (TP-py) has been synthesised and evaluated for its one- and two-photon absorption (2PA) induced-fluorescence properties under biological conditions. Interestingly, these compounds ore only weakly fluorescent in water whereas their fluorescence emissions are strongly restored (exaltation factors of 20-100) upon binding to double-stranded DNA. Additional measurements in glycerol indicate that the fluorescence increases are the result of immobilisation of the dyes in the DNA matrix, which inhibits rotational de-excitation modes. This particular feature is especially remarkable in the case of the bis and tris derivatives (TP-2py, TP-3py), which each display a high affinity (K-d similar to mu M) for dsDNA. TPIF measurements have shown that TP-2py and TP-3py each have a large 2PA cross section (8 up to 700 GM) both in glycerol and in the presence of DNA, which ranks them amongst the best 2PA biological fluorophores. Finally, one- and two-photon confocal imaging in cells revealed that these compounds perform red staining (lambda(em) = 660-680 nm) of nuclear DNA with excellent contrast. The remarkable optical properties of the TP-py series, combined with their high photostability and their easy synthetic access, make these compounds extremely attractive for use in confocal and 2PA microscopy.