Binding radioligand

Typically, filtration is used to separate bound from free using 96-well plates in a total volume of 200 iL, but SPA technology using yttrium silicate wheatgerm agglutinin-coated beads in 384-well plates and a total volume of 60 pL has also been used [43]. 

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There should be specific, saturable binding to the receptor, accompanied by pharmacological characteristics appropriate to the functional effects, demonstrable using a radioactive, eg, tritium or iodine-125, ligand to label the receptor. Radioligand binding assays (1,6) have become a significant means by which to identify and characterize receptors and enzymes (see Immunoassays Radioactive tracers). Isolation of the receptor or expression of the receptor in another cell, eg, an oocyte can be used to confirm the existence of a discrete entity. 

CB, was identified by radioligand binding techniques and subsequently cloned. A second receptor subclass, CB2, has been identified in human spleen and also has been cloned (34). Table 5 Hsts cannabinoid receptor ligands. 

Direct quantitation of receptor concentrations and dmg—receptor interactions is possible by a variety of techniques, including fluorescence, nmr, and radioligand binding. The last is particularly versatile and has been appHed both to sophisticated receptor quantitation and to dmg screening and discovery protocols (50,51). The use of high specific activity, frequendy pH]- or p lj-labeled, dmgs bound to cmde or purified cellular materials, to whole cells, or to tissue shces, permits the determination not only of dmg—receptor saturation curves, but also of the receptor number, dmg affinity, and association and dissociation kinetics either direcdy or by competition. Complete theoretical and experimental details are available (50,51). 

Radioligand binding to receptor dimers demonstrating co-operative behavior (3.13.1)... 

Radioligand Binding to Receptor Dimers Demonstrating Cooperative Behavior... 

FIGURE 4.12 Effect of alcuronium on the binding of [3H] methyl-QNB (filled circles) and [3H] atropine (open circles) on muscarinic receptors. Ordinates are percentage of initial radioligand binding. Alcuronium decreases the binding of [3H] methyl-QNB and increases the binding of [3H] atropine. Data redrawn from [7]. 

The Langmuir adsorption isotherm for radioligand binding [A ] to a receptor to form a radioligand-receptor complex [A R] can be rewritten in terms of one where it is not assumed that receptor binding produces a negligible effect on the free concentration of ligand ([A free]) ... 

Ehlert, F. J. (1988). Estimation of the affinities of allosteric ligands using radioligand binding and pharmacological null methods. Mol. Pharmacol. 33 187-194. 

Table 11.15 shows the mean of four estimates of the affinity of an antagonist measured with radioligand binding... 

Ansell, RJ Mosbach, K, Magnetic Molecularly Imprinted Polymer Beads for Drug Radioligand Binding Assay, Analyst 123, 1611, 1998. 

In practice, it is not often possible to directly measure par except in radioligand binding experiments. In many experiments it is the relationship between agonist concentration [A] and percentage maximum response (y) which is measured (a dose-response curve) and the Hill plot is made by plotting... 

Nearly all the derivatives of 2,5-DMA exhibited radioligand binding charae-teristies at 5-HT2 serotonin reeeptors that were consistent with those of serotonin and other tryptamine agonists. It has been demonstrated... 

Titeler. M. Lyon, R.A. and Gleimon, R.A. Radioligand binding evidence implicates the brain 5-HT2 receptor as a site of action for LSD and phenylisopropyl amine hallucinogens. Psychopharmacol 94 213-216,... 

McPherson, G.A. A practical computer-based approach to the analysis of radioligand binding experiments. Comput Programs Biomed 17 107-114, 1983. 

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