Errors Introduced in Radioligand Binding Studies Due to Displaceable, Cation Dependent, [3H]prazosin Binding to Glass-Fibre Filters and Glass Surfaces
| Autor: | D. M. J. Veenstra, M. J. Krielaart, Frans P. Nijkamp, K.J.H. van Buuren |
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| Rok vydání: | 1993 |
| Předmět: |
Male
Guinea Pigs Glass fiber Population Analytical chemistry Radioligand Assay Cations Prazosin medicine Animals Binding site Phentolamine education Cerebral Cortex Pharmacology education.field_of_study Binding Sites Chemistry Dissociation constant Membrane Radioligand binding Biophysics Glass Saturation (chemistry) Filtration medicine.drug |
| Zdroj: | Journal of Receptor Research. 13:801-814 |
| ISSN: | 0197-5110 |
| DOI: | 10.3109/10799899309073694 |
| Popis: | [3H]prazosin not only specifically and homogeneously labels alpha 1-adrenoceptors, but also binds to glass surfaces and non-linearly to the glass-fibre filters, commonly used in radioligand binding experiments. Binding to filters can be modulated by unlabeled alpha-adrenergic compounds and cations. If no correction is applied for displaceable filter binding, analysis of [3H]prazosin binding experiments leads to erroneous results. Analysis of [3H]prazosin saturation experiments on guinea-pig cerebral cortex membranes with correction for filter binding before the non-linear fit procedure indicated that [3H]prazosin labels a homogeneous population of alpha 1-adrenoceptors (Rtot: 8.33 fmol.mg-1 wet tissue) with a dissociation constant of 1.28 x 10(-10) M. However, analysis of the same data after correction for non-specific binding, (determined in parallel experiments by adding 10 microM phentolamine to the incubation medium) resulted in a best fit to a model in which [3H]prazosin labels two alpha 1-adrenoceptor subpopulations (R1: 15.0 fmol.mg-1 and R2: 14.6 fmol.mg-1 wet tissue) with dissociation constants of respectively 1.78 x 10(-10) and 5.63 x 10(-9) M. The discrepancy between the two methods of analysis is due to displacement of the radioligand from the filters by phentolamine. Prazosin and oxymetazoline are also able to displace filter-bound [3H]prazosin. The extent to which displaceable filter binding distorts the proper results depends on the actual magnitude of the error and also on the method of analysis. |
| Databáze: | OpenAIRE |
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