Modes of Determining β‐Adrenoceptor Number in Human Mononuclear Leucocytes

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Abstract
Inhibition of total125I‐ICYP binding to intact human mononuclear leucocytes at 32° by propranolol and ($pL)CGP‐12177 was biphasic. The high affinity component of125I‐ICYP binding, representing approximately 30% of total, was stereospecific, while the low affinity binding site was inhibited without sterospecificity. (‐)Isoproterenol inhibited the high affinity component of125I‐ICYP binding only, with low affinity. By performing binding studies in intact cells at 4° or in broken cell preparations at 37°, the fraction of total125I‐ICYP binding representing specific binding was increased, and agonist affinity was high. Inhibition of3H‐CGP‐12177 binding to intact cells at 32° demonstrated a high fraction of specific binding and high agonist affinity. Computer‐assisted analysis of total radioligand binding determined over a broad concentration range revealed two populations of saturable125I‐ICYP binding sites in intact cells as well as in broken cell preparations, while3H‐CGP‐12177 binding demonstrated only one saturable binding site. The number of high affinity125I‐ICYP binding sites was comparable to the number of saturable3H‐CGP‐12177 binding sites. Receptor numbers determined by analysis of total radioligand binding were comparable to receptor numbers determined by subtraction of non‐specific binding, determined in the presence of a high concentration of competing ligand. Analysis of total radioligand binding was found to be a better procedure because it eliminates the use of an arbitrary concentration of unlabelled ligand and improves the accuracy of the assay.