Radio-ligand binding assay

The riboflavin binding protein that occurs in eggs has been exploited for the radio-ligand binding assay of riboflavin. Because binding to the protein quenches the native fluorescence of riboflavin, it can be exploited for a direct titrimetric fluorescence assay of the vitamin in urine and other biological samples (Kodentsova et al., 1995). 

Kll. Korenman, S. G., Radio-ligand binding assay of specific estrogens using a soluble uterine macromolecule. J. Clin. Endocrinol. 28, 127-130 (1968). 

Optimization of molecularly imprinted polymers for radio-ligand binding assays... 

Mayes, A. Lowe, C.R. Optimization of molecularly imprinted polymers for radio-ligand binding assays. Royal Society of Chemistry Cambridge, UK, 1998 Vol. 25, 28-36. Andersson, L.I. Muller, R. Vlatakis, G. Mosbach, K. Mimics of the binding sites of opioid receptors obtained by molecular imprinting of enkephalin and morphine. Proc. Natl. Acad. Sci. USA 1995, 92, 4788 792. 

Towards this end, studies were carried out which included the radio-labelling of the somatostatin receptor ligand DOTA-Tyr -TATE (1,4,7,10-tetraazacyclotetradecane-N,N, N",N" -tetraacetic acid tyrosineS-octreotate, or DOTATATE) with both Lu and I, and the quality control of the resulting complexes. In vitro studies of the biological affinity of the Lu-DOTATATE and I-DOTATATE for the somatostatin membrane receptor were carried out with the help of competition and saturation binding assays. In vivo studies of the biodistribution of Lu-DOTATATE and I-DOTATA TL in animals, either alone or in competition, were also carried out. The radiotherapeutic effect of "Lu-DOTATATE was evaluated by cytometry measurements. Estimation of absorbed doses of Lu-DOTATATE was carried out by mathematical modelling. 

Desialylated human chorionic gonadotrophin was found to be twice as effective as the native hormone in competing for the binding site of receptors in rat testis. However, it has been pointed out that caution must be exercised when radio-ligand receptors are used to assay preparations of human chorionic gonadotrophin of different sialic acid contents. 

Development of these assays can be difficult. A radio-labeled ligand that binds to the target receptor with high affinity is required, along with sufficient receptor and sufficient radioactivity to cause the SPA bead to signal. These aspects may limit the ability of a specific assay to be miniaturized into a 1536-well plate format. However, for some receptors and ligands, it has proven an effective way to screen. 

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... 

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