that the concentration of the receptor (Ttot) was «Kd and the bound radioligand was quantified after filtration as a separation step. For the test compounds (+)-butaclamol, chlorpromazine and (S)-sulpiride, the resulting Ki values determined by conventional [3H]spiperone binding assays were 2.5- to 10-fold lower than those established by MS binding studies (Table 7.2) . The reasons for the, partly considerable, deviations are still unknown. However, they are obviously not the result ofthe mass spectrometric quantitation as the identically conducted control experiment quantifying nonbound [3H]spiperone using (S)-sulpiride as competitor had yielded results similar to the MS binding assay. The main cause is presumably the large amount of membrane fraction required to obtain a sufficient concentration of binding sites for the marker. Particularly, the lipophilic test compounds (+)-butaclamol and chlorpormazine might show a high amount of nonspecific binding to the membrane fraction resulting in ligand depletion which would easily explain the rise of IC50 and Ki values, respectively. Of course this problem could be effortlessly solved by employing a more appropriate receptor source with a higher receptor density, e.g. a membrane fraction of a heterolo-gous expression system, or by using a more powerful mass spectrometer.
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