Table 1.
Characteristics of membranes derived from cells transiently expressing prostanoid receptors and of the competition radioligand binding assays using them
| |
Membranes |
Assay |
||||||||
|---|---|---|---|---|---|---|---|---|---|---|
| Receptor | Vector | Bmax (pmol mg−1) | Kd (nM) | Protein (μg per well) | Assay type | Radioligand | Specific activity (TBq mmol−1) | [radioligand] (nM) | n.s.b. ligand | [n.s.b.] (μM) |
| DP |
pcDNA3 |
12.3 |
11.9 |
12.0 |
SPA |
[3H]-PGD2 |
5.96 |
1 |
PGD2 |
10 |
| EP1 |
SFV-1 |
3 |
12 |
11.3 |
Filtration |
[3H]-PGE2 |
6.07 |
10 |
PGE2 |
100 |
| EP2 |
SFV-1 |
4.3 |
19 |
7.8 |
SPA |
[3H]-PGE2 |
6.07 |
10 |
PGE2 |
100 |
| EP3I |
SFV-1 |
30 |
6 |
2.3 |
SPA |
[3H]-PGE2 |
6.07 |
3 |
PGE2 |
100 |
| EP4 |
SFV-1 |
1.1–8.8 |
3 |
0.7–2.4 |
SPA |
[3H]-PGE2 |
6.07 |
3 |
PGE2 |
100 |
| FP |
pcDNA3 |
12 |
8.2 |
18.0 |
SPA |
[3H]-PGF2α |
7.88 |
2.1 |
PGF2α |
10 |
| IP |
pcDNA3 |
6.5 |
13 |
15.0 |
SPA |
[3H]-iloprost |
0.63 |
15 |
iloprost |
1 |
| TP | pcDNA3 | 6.4 | 7 | 15.0 | SPA | [3H]-SQ29,548 | 1.80 | 6 | U-46619 | 1 |
Bmax: estimated receptor expression (pmol mg−1); Kd: the radioligand dissociation-binding constant (affinity) determined by saturation analysis (nM); protein: amount of protein per well for each assay (μg well−1); [radioligand]: concentration of radioligand (nM); n.s.b. ligand: the nonradiolabelled ligand used to displace radioligand from the receptors and therefore determine nonspecific binding; [n.s.b.]: concentration of n.s.b. ligand (μM).