Tissue Factor Pathway Inhibitor (TFPI) represents the main physiologic inhibitor of the coagulation start. We detected TFPI together with Tissue Factor in the a-granules of resting platelets. mRNA analyses indicated that platelets exhibit high transcript levels for TFPI-a, but very low levels for TFPI-b. After stimulation with thrombin and collagen, TFPI was recovered in the platelet releasate. Following conjugate formation between the activated platelets and polymorphonuclear neutrophils (PMN), substantial TFPI degradation was noted. This decomposition of TFPI could be prevented by specific inhibitors against neutrophil elastase (NE) and cathepsin G (CG), but not by suppression of matrix metalloproteinases (MMP). In addition, the formation of factor Xa by platelet-neutrophil suspensions was prevented by inhibiting NE or CG, but not by obstructing the MMP. The TFPI degradation by PMN was also inhibited by pre-coating isolated PMN with anti-PSGL-1 or anti-CD18-antibodies. Our results suggest, that upon interaction of activated platelets with PMN a microenvironment is formed, which allows the efficient degradation of TFPI by the PMN-associated serine proteases NE and CG, thus triggering the coagulation start. This mechanism might be of general importance in pathologies like sepsis, arterial and venous thrombosis and myocardial infarction.