ORIGINAL ARTICLESThe intrinsic coagulation activity assayStief, Thomas W; Otto, Stefanie; Renz, HaraldAuthor Information Department of Clinical Chemistry — Central Laboratory, University Hospital Giessen & Marburg, Germany Received 6 February, 2006 Accepted 13 March, 2006 Correspondence and requests for reprints to T. W. Stief, MD, Priv.-Doz., Department of Clinical Chemistry, University Hospital, D-35033 Marburg, Germany fax: +49 6421 286 5594; e-mail: [email protected] Blood Coagulation & Fibrinolysis: July 2006 - Volume 17 - Issue 5 - p 369-378 doi: 10.1097/01.mbc.0000233367.95733.d8 Buy Metrics Abstract A new assay for the contact-phase-mediated generation of thrombin activity has been developed – the intrinsic coagulation activity assay (INCA). Citrated plasma (50 μl) is incubated with 5 μl SiO2, 250 mmol/l CaCl2 in polystyrole flat-bottom wells. After exactly 4 and 5 min (37°C) coagulation reaction times (INCA-4 and INCA-5), 100 μl of 2.5 mol/l arginine, pH 8.6, is added to inhibit hemostasis activation in the important ascending part of the thrombin generation curve and to depolymerize fibrin. After 20 min, 50 μl of 1 mmol/l (final concentration 0.24 mmol/l) chromogenic thrombin substrate CHG-Ala-Arg-pNA in 1.25 mol/l arginine, pH 8.7, is added. The increase in absorbance is determined at 405 nm using a microtiterplate photometer. The assay is calibrated against 1 IU/ml thrombin. The normal thrombin activity range of INCA-4 (main value) or INCA-5 (control value) is 100 ± 30% of normal (mean value ± 1 SD; 100% = 0.5 IU/ml for INCA-4 and 1.9 IU/ml for INCA-5). With the INCA the normal range of intrinsic hemostasis is reflected, low-molecular-weight heparins can be monitored, the plasma matrix is not changed significantly, and the assay results are a percentage of normal generated thrombin activity and not coagulation seconds. © 2006 Lippincott Williams & Wilkins, Inc.