Ligand binding assay surface plasmon resonance

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. 

Figure 10 Surface plasmon resonance analysis. A surface displaying ligand is examined under conditions of flow. Injection of lectin results in an increase in signal observed in the association phase of the curve. At equilibrium the surface achieves maximal binding. Measurement of the dissociation can yield off-rates. Competing ligands in solution can also be assayed.

The individual sequences can be tested for their affinity for the target protein by various methods like filter binding assay, gel-shift, and surface plasmon resonance. The individual sequences from each class can be synthetically prepared from DNA synthesis facilities and tested for their ligand-binding capability. A representative binding curve from which the binding affinity can be estimated is shown in Fig. 3. 

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