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Fig. 7.7 Representative binding curve obtained by nonlinear regression from a competitive MS binding assay for dopamine D1 receptors, in which (+)-butaclamol competes with SCH 23390 as marker. The points describe nonbound SCH 23390 quantified by LC-ESI-MS/MS. Data reflect means (+s) from binding samples, each performed in quadruplicate.

Table 7.1 Affinities (mean + SEM, n = 3) for dopamine antagonists at receptors obtained by MS-binding assays and radioligand binding assays, respectively [61].

SCH 23390 [3H]SCH 23390

Chlorpromazine 1700 + 40 620 + 10 300 + 40

Pimozide 13 000 + 700 4700 + 200 2500 + 500 SKF 83566 5.9 + 1.0 1.9 + 0.3 2.7 + 0.6

(S)-Sulpiride >10000 >10000 >10000

Trifluoperazine 1300 + 100 460 + 40 215 + 20

in the same way and their affinity constants calculated from the resulting competition curves (Table 7.1, Fig. 7.4).

To validate the results, the test compounds were examined in radioligand binding assays under comparable conditions. In this case, however, according to the common procedure in conventional radioligand binding assays, a target concentration [Ttot ] « Kd was employed, and [3H]SCH 23390 bound to the target was quantified after filtration (Table 7.1). The Ki values determined [according to Eq. (7), Section 7.3.1) in the MS binding assays tend to be slightly higher than those determined in radioligand binding assays. However, most importantly the ranking of the Ki values is in good accordance for both test systems [61].

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