Radioligand binding studies analysis

The MIPs have also been utilized as the recognition elements in pseudoimmunoassays. " In this approach, MIPs are substituted for antibodies to quantify the amount of analyte in a biological sample, such as blood plasma. Most MIP immunoassays are competitive binding studies in which a radio- or fluorescent-labeled analyte is added to a mixture of the MIP and imlabeled analyte. After equilibrium is reached, some fraction of the labeled species is bound to the polymer surface and thus can be separated from the supernatant. The supernatant is then analyzed via scintillation or fluorescence techniques to determine the concentration of the original unlabeled analyte. Mosbach et al. have demonstrated that MIP-based immunoassays can rival the selectivity of antibody-based assays. Imprinted polymers for the opioid receptor ligands enkephalin and morphine were prepared and showed submicromolar (pM) level selectivity in a radioligand competition assay in aqueous buffers. The analysis... 

The thermodynamic analyses most often used, particularly fhe van t Hoff mefhod, require that measurements be made at steady-state conditions. In the case of radioligand binding determination of equilibrium constants, fhe time required for fhe protein-ligand interaction to reach steady state depends on fhe incubation temperature, and, therefore, the equilibrium constant must be determined for each temperature studied. For the most accurate results, fhe determination needs to be made at more than two temperatures in order to detect non-linearity. The integrated form of the van t Hoff equation takes fhe simple form fhat is commonly used only if ATT and AS° for the interaction are not temperature dependent otherwise, non-linearity in the van t Hoff plot can arise. Meaningful information can StiU be obtained in such cases, but more complex analysis is required. 

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