Site | 3H-Ligand | Ligand | Nonspecific Binding | Tissue | No. Washes | Buffer1-a | Incubation | Filter Soak1-b |
---|---|---|---|---|---|---|---|---|
nM | mg wet wt | ×/volumes | h/°C | min | ||||
DAT | CFT | 2.0 | 100 μM (−)-cocaine | 4.0 Caudate/putamen1-c | 1 /10 | 1) 50 mM Tris | 2 /4 | 40/0.1% BSA |
2 /40 | 2) 50 mM Tris | |||||||
100 mM NaCl | ||||||||
NET | Nisoxetine | 3.0 | 1.0 μM mazindol | 8.0 Cerebellum | 3 /30 | 1) 50 mM Tris | 4 /4 | 240/0.3% polyethylenimine |
120 mM NaCl | ||||||||
5 mM KCl | ||||||||
2) 50 mM Tris | ||||||||
300 mM NaCl | ||||||||
5 mM KCl | ||||||||
SERT | Paroxetine | 0.10 | 1.0 μM sertraline | 3.0 Frontal cortex | 2 /50 ml | 50 mM Tris | 1 /25 | 60/0.03% polyethylenimine |
120 mM NaCl | ||||||||
5 mM KCl |
NET, no epinephrine transporter; SERT, serotonin transporter.
↵1-a 1, homogenization buffer; 2, incubation buffer. Homogenization and incubation buffers were identical for paroxetine.
↵1-b Filters were presoaked in assay buffer unless otherwise indicated.
↵1-c Approximately equal milligram wet weight amounts of caudate and putamen tissue were used.