Binding kinetics problems associated with

Lanthanide chelate-dyed polystyrene particles containing carboxylic acid groups have been covalently coated with antibodies, streptavidin [95], and nucleic acids [96] for different biomolecular-binding assays. Particulate labels can be coated with antibodies and used as such, but the obvious steric and kinetic problems associated with the large molecular size can be particularly avoided by indirect detection of the bound antibodies using, e.g., biotin-streptavidin interaction [97,... 

Here we describe studies of the interaction of interleukin-6 (IL-6) with a soluble form of its cell surface receptor (sIL-6R). A procedure utilising a competition approach is presented which allows the determination of the equilibrium constant in solution thus avoiding any potential problems associated with deviation in kinetic characteristics upon surface immobilisation. In addition, binding characteristics of stable monomeric and dimeric forms of IL-6 are presented to demonstrate both the drastic influence of solute multivalency on kinetic and equilibrium properties and the importance of auxiliary techniques such as analytical ultracentrifugation for the interpretation of SPR data. 

In the kinetic studies of the adsorption process, the mass transport of the analyte to the binding sites is an important parameter to account for. Several theoretical descriptions of the chromatographic process are proposed to overcome this difficulty. Many complementary experiments are now needed to ascertain the kinetic measurements. Similar problems are found in the applications of the surface plasmon resonance technology (SPR) for association rate constant measurements. In both techniques the adsorption studies are carried out in a flow system, on surfaces with immobilized ligands. The role of the external diffusion limitations in the analysis of SPR assays has often been mentioned, and the technique is yet considered as giving an estimate of the adsorption rate constant. It is thus important to correlate the SPR data with results obtained from independent experiments, such as those from chromatographic measurements. 

Tb complex in this system, [Tb (19)], is octaden-tate, thus removing the problem of self-association as observed in the previous systems. Titrations monitoring the Tb centered emission in aqueous buffer solutions (Phosphate Buffered Saline (PBS) and 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) both at pH 7.4, where the isophthalate is fully deprotonated), indicate successful formation of [Eu2(18)] [Tb(19)] with comparable binding constants for each of the buffer systems of K = 225 600 M- (PBS) and 226500 (HEPES). These results highlight the ternary complex formation approach to generating heterometallic 4f systems through carefiil matching of interactions between kinetically stable complexes. 

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