Radioligand competitive binding methodology for the evaluation of platelet-activating factor (PAF) and PAF-receptor antagonism using intact canine platelets.
High-affinity, stereoselective, and ligand-selective specific binding of 3H-labeled platelet-activating factor (PAF) to its receptor on the dog platelet is reproducible over wide mass and concentration ranges of [3H]PAF. The [3H]PAF specific binding can be competitively inhibited by low picogram amounts of nonlabeled PAF. These observations have led to the formulation of radioligand competitive binding methodology for the detection and estimation of PAF in a biological lipid sample and the quantitative evaluation of PAF-receptor antagonism. The methodology is predicated upon correlation between the ability of a PAF analog/biological lipid sample/(synthetic) substance to inhibit [3H]PAF specific binding to the washed canine platelet and the known inhibition of [3H]PAF specific binding by standard, nonradioactive PAF. Application of this methodology to lipid extracts of human saliva has uncovered the finding that subjects with upper respiratory infection and chronic allergies have high saliva PAF contents. Pharmacologically active antiallergy agents known to inhibit PAF-induced pathology in animal models of disease were demonstrated, with the methology advanced, to act as PAF-receptor antagonists, and their potencies were quantified. These investigations indicate that the system proposed, in its ease, economy, sensitivity, specificity and capacity, has practical value for detecting and estimating PAF in biological lipid extracts and for evaluating PAF-receptor antagonism.
Janero
D R
DR
Department of Pharmacology and Chemotherapy, Roche Research Center, Hoffmann-La Roche Inc., Nutley, New Jersey 07110.
Burghardt
B
B
Burghardt
C
C
D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D001792:Blood Platelets / Q000187:drug effects*; D004285:Dogs; D005069:Evaluation Studies as Topic; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D010743:Phospholipids / Q000097:blood; D010972:Platelet Activating Factor / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010974:Platelet Aggregation / Q000187:drug effects; D010980:Platelet Membrane Glycoproteins; D011869:Radioligand Assay; D011956:Receptors, Cell Surface / Q000187:drug effects*; D043562:Receptors, G-Protein-Coupled; D012463:Saliva / Q000378:metabolism; D013997:Time Factors
D010743:Phospholipids; D010972:Platelet Activating Factor; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; C039351:platelet activating factor receptor