Platelet-activating factor binding to human platelet membranes.
Previously reported methods for quantifying platelet-activating factor (PAF) binding to rabbit platelet membranes were modified for studies of PAF binding to human platelet membranes. The membranes were prepared by the "glycerol lysis" method and PAF binding was quantified by using polyethylene glycol precipitation to recover membrane-bound PAF. Optimal PAF binding required buffers containing 3 to 10 mm KCl and either 5 to 10 mM MgCl2 or 5 to 10 mM CaCl2. NaCl was not as effective as KCl and concentrations of NaCl greater than 3 mM strongly inhibited PAF binding. Maximal binding occurred after incubation for 60 min at 0 degree C and was reversed by the addition of excess unlabeled PAF. PAF binding was saturable. Scatchard analysis of PAF binding to 50 micrograms of membrane protein revealed 10.3 +/- 1.7 x 10(11) receptors per milligram of membrane protein and the receptors had a Kd of 7.6 +/- 1.9 nM. The calculated receptor number, binding affinity, and specificity of binding are similar to those previously calculated for PAF binding to intact human platelets, suggesting that the membrane binding site for PAF is the PAF receptor.
D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D002122:Calcium Chloride / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D006801:Humans; D008274:Magnesium / Q000494:pharmacology; D015636:Magnesium Chloride; D010972:Platelet Activating Factor / Q000378:metabolism*; D010980:Platelet Membrane Glycoproteins; D011189:Potassium Chloride / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D011817:Rabbits; D011956:Receptors, Cell Surface / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled; D012965:Sodium Chloride / Q000494:pharmacology
D010972:Platelet Activating Factor; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; C039351:platelet activating factor receptor; D015636:Magnesium Chloride; D012965:Sodium Chloride; D011189:Potassium Chloride; D008274:Magnesium; D002122:Calcium Chloride