USE OF MODIFIED FUNCTIONAL ASSAYS FOR ACTIVATED PROTEIN-C RESISTANCE IN PATIENTS WITH BASALLY PROLONGED APTT

Inherited resistance to activated protein C (APCr) is currently recogn ized as the most prevalent cause underlying venous thrombophilia, with an estimated prevalence around 20% in thrombotic patients and around 1.8-7% in the general population. A correct laboratory diagnosis of AP Cr is therefore essential. Two different diagnostic approaches are at present at our disposal: the semi-quantitative plasma test based on th e measurement of two aPTTs (in the presence and absence of activated p rotein C), and the detection of the factor V Arg(506) Gin mutation by DNA analysis. In this study we firstly evaluated sensitivity, specific ity and diagnostic efficiency of an aPTT-based plasma clotting test (C hromogenix, Sweden) versus DNA analysis: then. since the APC resistanc e test is invalidated by a basally prolonged aPTT (i.e, during warfari n and heparin therapy or in patients with clotting factor deficiencies or in the presence of a lupus anticoagulant), patient plasmas were co nveniently diluted in factor V deficient plasma in order to correct cl otting factor abnormalities. Nevertheless, patients with a LA and an a PTT ratio range 1.8-3.17 were still all misclassified. We obtained com et diagnoses in LA positive patients by preincubating plasmas with a m ixture of phospholipids; therefore we decided to perform a double modi fied clotting test adding a mixture of platelet derived phospholipids to samples previously diluted in factor V deficient plasma. The perfor mance characteristics of this novel method with a different aPTT reage nt (Behring Germany) were also evaluated. With this double modified te st all patients were correctly classified as negative or positive for factor V mutation in agreement with DNA analysis, irrespectfully of th e basal aPTT value and the aPTT reagent employed. We propose this modi fied version of the APCr clotting test as an easily reproducible, reli able, very sensitive and specific screening test which possibly reduce s the need for DNA analysis.