Ruthenium Complex Labels

d⁶ metal ions such as Ru²⁺, Os²⁺, Re⁺ form highly fluorescent complexes with chelating ligands such as bipyridines and phenanthrolines. Additional ligands such as phosphines, nitriles, CO or pyridines can be used to fine-tune their properties and/or for labelling purposes.

The labels obtained from these compounds are remarkably stable both chemically and photochemically and possess unique properties, such as long fluorescence lifetimes (from hundreds of nanoseconds to several microseconds) and very large Stokes shifts.

The long lived fluorescence signal of these labels can be easily distinguished from the short lived emission from cells and biomolecules, while minimizing dye self absorption. In addition, for some of these fluorophores, it is possible to use electricity rather than light to elicit a fluorescent signal.

This phenomenon, known as Electrochemiluminescence (ECL) can be exploited to achieve extremely high signal-to-noise ratios. These probes can be used in fluorescence polarization assays, time-resolved and ECL immunoassays and imaging applications.1-4

Absorption and Emission Spectra of [Tris(bathophenanthrolinedisulfonate)Ru(II)] tetrasodium salt hexahydrate

References:

1. Bard, A. J.(Ed.) Electrogenerated Chemiluminescence, Marcel Dekker, New York, 2004.

2. Richter, M. M. Chem. Rev., 2004, 104, 3003.

3. Miao, W. Chem. Rev. 2008, 108, 2506.

4. Xu, X. N. and Zu, Y. In New Frontiers in Ultrasensitive Bioanalysis, Xu, X. N. (Ed.), Wiley, New York, 2007, pp 235-268.

Neutral and dianionic ruthenium bathophenanthrolinedisulfonated complexes: a route to enhance chemiluminescence performance in aqueous media

We reported5 the strong enhancement of ECL intensity and duration in neutral and dianionic Ru(II) complexes bearing mixed 2,2’-bipyridine (bpy) and bathophenanthrolinedisulfonate (BPS) ligands.
In aqueous conditions, using the tripropylamine-assisted method and applying a constant potential, we observed for Ru(BPS)(bpy)2 a remarkable 26-fold ECL integrated intensity increase with respect to Ru(bpy)32+. The results herein obtained reveal that the cause of reduced ECL intensity of [Ru(BPS)3]4- and [Ru(BPS)2(bpy)]2- can be ascribed to surface effects related to the interaction of the oxidized complex with the electrode.
The expected reduction or absence of electrostatic interactions with biomolecules, together with the strongly enhanced performance, makes the zwitterionic Ru(BPS)(bpy)2 complex a highly promising candidate for the development of very efficient ECL labels for ultrasensitive bio-assays and functional imaging applications.

5. Della Ciana, L., Zanarini, S, Perciaccante, R., Marzocchi, E. and Valenti, G. Ru(bpy)32+ Neutral and Dianionic Ru(II) Bathophenanthroline disulfonate Complexes: a Route to Enhance Chemiluminescence Performance in Aqueous Media, J. Phys. Chem. C, , (2010) 114 (8), pp 3653–3658.

Ru(bpy)32+ covalently doped silica Nanoparticles as multicenter tunable structures for electrochemiluminescence (ECL) amplification

We investigated6 the ECL of doped silica nanoparticles (DSNPs), prepared by a reverse microemulsion method that leads to covalent incorporation of the Ru(bpy)32+, ion in acetonitrile and aqueous buffers. The emission was produced for the first time by cation-anion direct annihilation, and the position of ECL maxima indirectly allowed estimation of the E1/2,IOx and E1/2IRed potentials of Ru(bpy)32+ inside the DNSP. Thiol terminated DSNPs were self-assembled on gold substrates, forming compact and stable monolayers which mimic probe-target assays with DSNPs as labels.

The ECL intensity obtained by such functionalized substrates in aqueous media, using tripropylamine (TPrA) as coreactant, was surprisingly increased with respect to direct electrochemical oxidation because of the ability of oxidized TPrA to diffuse within the DSNPs structure and reach a higher number of emitting units with respect to direct electron tunneling. The experimental results were explained by proposing a basic physical-chemical model, which supports evaluation of the number of redox-active centers per nanoparticle. In the model the contrasting effects of increased luminescence quantum yield and decreased diffusion coefficient with respect to free (i.e., not bound within the silica structure) emitting molecules were taken into account. This allows, in principle, optimizing the ECL emission intensity as a function of DSNP size, doping material, charge, doping level, supporting electrolyte, electrode material, and solvent.

In conclusion, this study has provided a more than 1000-fold increase of the ECL signal of a chemically and electrochemically stable DSNP compared to that of a single
dye, suggesting that use of this kind of nanostructures as luminescent labels represents a very promising system for ultrasensitive bioanalysis.

6. Zanarini, S, Rampazzo, E., Della Ciana, L., Marcaccio, M., Marzocchi, E., Montalti, M., Paolucci, F. and Prodi, L. Ru(bpy)32+ Covalently Doped Silica Nanoparticles as Multicenter Tunable Structures for Electrochemiluminescence Amplification, J. Am. Chem. Soc. (2009), 131 (6), 2260-2267.