Platelet-activating factor (PAF) is released in IgE-mediated allergic diseases. The normal level, the method of its determination and its clinical importance are subject of controversy. We hypothesized that a functional assay could help to better analyze the actual concentrations in vivo because PAF may be released locally and is short-lived. An assay to detect PAF by the desensitized state of human platelets exposed to PAF in vitro or ex vivo was developed: We analyzed the synergistic platelet response to dual agonist stimulation at extraordinarily low doses (collagen 0.10 µg/ml and PAF 2.5 × 10^––8M) in aggregation and release reaction and its absence after previous exposure to PAF at concentrations between 5 × 10^––9 and 5 × 10^––11M in vitro. The same test was then applied to examine the platelets from patients with IgE-mediated allergic asthma before and after inhalation of the specific allergen (inhalative provocation test; a reduction of the FEV₁ by > 15% was considered positive). The lack of a synergistic response to collagen with PAF was found after preincubation of the platelets with 5 × 10^––9M PAF and a reduction of ± 50% with 5 × 10^––10M in vitro. A significant reduction of the aggregation response (––56 ± 18%) and of the release of Β-thromboglobulin (––75 ± 24%) was found in 6 patients with a positive inhalative provocation test but not in 3 patients with a negative response. We conclude that PAF is released in asthma in man at levels of ∼ 10^––10 M after challenge with the specific allergen and induces a desensitization of the platelets to PAF. This mechanism may explain the platelet activation in asthma observed earlier and our assay may provide a useful diagnostic tool to document the release of PAF in vivo.