Lanthanides, chelates

Vereb, G., Jares-Erijman, E., Selvin, P. R. and Jovin, T. M. (1998). Temporally and spectrally resolved imaging microscopy of lanthanide chelates. Biophys. J. 74, 2210-22. 

Different lanthanide metals also produce different emission spectrums and different intensities of luminescence at their emission maximums. Therefore, the relative sensitivity of time-resolved fluorescence also is dependent on the particular lanthanide element complexed in the chelate. The most popular metals along with the order of brightness for lanthanide chelate fluorescence are europium(III) > terbium(III) > samarium(III) > dysprosium(III). For instance, Huhtinen et al. (2005) found that lanthanide chelate nanoparticles used in the detection of human prostate antigen produced relative signals for detection using europium, terbium, samarium, and dysprosium of approximately 1.0 0.67 0.16 0.01, respectively. The emission... 

Lanthanide chelates also can be used in FRET applications with other fluorescent probes and labels (Figure 9.51). In this application, the time-resolved (TR) nature of lanthanide luminescent measurements can be combined with the ability to tune the emission characteristics through energy transfer to an organic fluor (Comley, 2006). TR-FRET, as it is called, is a powerful method to develop rapid assays with low background fluorescence and high sensitivity, which can equal the detection capability of enzyme assays (Selvin, 2000). 

Figure 9.51 Time-resolved FRET assay systems involve energy transfer between the lanthanide chelate and an organic dye that are brought together as two labeled molecules bind to an analyte. In this illustration, an antibody labeled with a lanthanide chelate is used along with a Cy5-labeled antibody to detect a protein target in solution. Excitation of the lanthanide label results in energy transfer and excitation of the cyanine dye only if they are held within close enough proximity to allow efficient FRET to occur. Under these conditions, excitation of the lanthanide chelate results in cyanine dye emission, which will not occur if the labeled antibodies have not bound to a target.

Organic fluorescent dyes with the appropriate spectral properties also can be paired with lanthanide chelates in FRET systems. For instance, many rhodamine dyes and the cyanine dye Cy5 have ideal excitation wavelengths for receiving energy from a nearby europium chelate. The LeadSeeker assay system from GE Healthcare incorporates various Cy5-labeled antibodies for developing specific analyte assays. In addition, if using a terbium chelate as the donor, then a Cy3 fluorescent dye can be used in assays as the acceptor. 

When used with europium or terbium ions, a carbostyril-based lanthanide chelate can be excited at 340 nm and provide sharp characteristic emission bands for transfer of energy to the appropriate acceptor fluor. Similar to the TMT chelator described previously, luminescence from terbium FRET signals well with Cy3 dyes and luminescence from europium can be used with APC or Cy5 dyes. Other fluorescent dyes that have similar excitation and emission ranges to these also can be used as acceptors in TR-FRET assays. For instance, terbium chelates can... 

Other fluorescent probes also may be used to label (strept)avidin molecules for detection of biotinylated targeting molecules. Chapter 9 reviews many additional fluorescent labels, such as quantum dots, lanthanide chelates, and cyanine dye derivatives, all of which may be used in similar protocols to create detection conjugates for (strept)avidin-biotin-based assays. 

Ge, P., and Selvin, P.R. (2004) Carbostyril derivatives as antenna molecules for luminescent lanthanide chelates. Bioconjugate Chem. 15, 1088-1094. 

Soini, E., and Kojola, H. (1983) Time-resolved fluorometer for lanthanide chelates—A new generation of monisotopic immunoassays. Clin. Chem. 29, 65-68. 

---