Lanthanide fluorophores

Keywords Amplification Fluorescent reporter Fluorophore FRET In vitro In vivo Labeling Lanthanide chelate Multiplexing Nanoparticle Quantum dot Transition metal complex... 

HTRF (homogeneous time-resolved fluorescence) LANCE (lanthanide chelate excitation) Cisbio International PerkinElmer Life Sciences Lanthanide TR-FRET using Eu3+ cryptate/chelate-donor fluorophore and cross-linked aUophycocyanin-acceptor fluorophore... 

LanthaScreen Invitrogen Lanthanide TR-FRET using Tb3+ chelate-donor fluorophore and fluorescein-acceptor fluorophore... 

For protein and DNA studies several methods of locating the lanthanide probe onto the chain have been employed. In one example the antenna fluorophore is covalently attached to one leg of the chelate of europium or terbium, which is further attached to the specific site through thiol reactive, e.g. maleimide, or amine reactive, e.g. A-hydroxysuccinimide, groups. A typical example is shown in (3.77). ... 

The use of europium chelates, with their unusually long fluorescence decay times, as labels for proteins and antibodies has provided techniques that are referred to as time-resolved fluoroimmunoassays (TRFIA). Fluorophores as labels for biomolecules will be the topic of Sect. 3. Nevertheless, TRFIAs always have to compete with ELISA (enzyme-linked immunosorbent assays) techniques, which are characterized by their great versatility and sensitivity through an enzyme-driven signal amplification. Numerous studies have been published over the past two decades which compare both analytical methods, e.g., with respect to the detection of influenza viruses or HIV-1 specific IgA antibodies [117,118]. Lanthanide luminescence detection is another new development, and Tb(III) complexes have been applied, for instance, as indicators for peroxidase-catalyzed dimerization products in ELISAs [119]. 

Time-resolved RET is capable of very sensitive detection of DNA hybridization. With a lanthanide chelate as the donor and an organic fluorophore like tetramethylrhodamin as the acceptor, time-resolved measurements can indicate the hybridization by strong changes in the intensity decay of the donor [186]. The development of new dyes for time-resolved RET with improved properties still is a major task [187,188]. But, so far, the detection of biomolecular interactions by time-resolved RET has not entered real applications in the DNA or protein array market. 

Cha et al. (1999) used a variant of FRET called LRET for lanthanide-based fluorescence energy transfer. In this technique (Selvin, 1996) the donor is terbium or europium which, in fact, is luminescent. There are several advantages of this technique over regular FRET. It has been found that terbium emits isotropically, which means that the uncertainty due to the dipole orientation is decreased to a maximum error of 10%. This error can be decreased even further if the anisotropy of the acceptor is also known. The second advantage is that the fluorescence decay has a time constant of about 1.5 ms, making it easily measurable with conventional recording techniques. The third advantage is that the emission of terbium is peaked and one can find fluorophores that emit in between peaks. This means that the fluorescence of the acceptor can be measured with little or no contamination from the donor. In addition, as the acceptor has a fast decay, any measurement of the acceptor fluorescence with decays comparable to the donor will exclude any possible direct... 

Owing to these imique properties, lanthanides have several advantages over traditional organic fluorophores, quantum dots, or other fluorescent species commonly used as sensors. These... 

---