. 2017 Jul 14;2(7):3616–3631. doi: 10.1021/acsomega.7b00451

Table 3. Conditions of the Radioligand Competitions Binding Assays at the NPY Receptor Subtypes.

	Y₁R	Y₂R	Y₄R	Y₅R
receptor source	MCF-7-Y₁ cells²³	CHO-hY₂R-G_qi5-mtAEQ cells²⁵	CHO-hY₄R-G_qi5-mtAEQ cells²⁶	HEC-1B-hY₅ cells²⁷
buffer	N-(2-hydroxyethyl)piperazine-N′-ethanesulfonic acid (HEPES) buffer containing 150 mM NaCl^a	sodium-free HEPES buffer^b	sodium-free HEPES buffer^b	HEPES buffer containing 150 mM NaCl^a
radioligand	compd 46	[³H]propionyl-pNPY	compd 45(11)	[³H]propionyl-pNPY
K_d	6.2 nM	1.4 nM	0.67 nM	4.8 nM
concentration	4 nM	1.0 nM	0.6 nM	4 nM
incubation period	90 min	90 min	90 min	120 min
separation	suction	suction	filtration	suction
scintillation cocktail	Optiphase Supermix	Optiphase Supermix	Rotiscint eco plus	Optiphase Supermix

Isotonic HEPES buffer pH 7.4 (150 mM NaCl, 10 mM HEPES, 25 mM NaHCO₃, 2.5 mM CaCl₂, 1.2 mM KH₂PO₄, 1.2 mM MgSO₄, 5 mM KCl).

Hypotonic, sodium-free HEPES buffer pH 7.4 (25 mM HEPES, 2.5 mM CaCl₂, 1 mM MgCl₂). Binding buffers were supplemented with BSA (1%) and bacitracin (0.1 mg/mL).