Commercial Thromboplastins Research Articles

PB1121 Laboratory INRs Determined with Commercial Thromboplastins in >450 Centres Before and after the Establishment of the IRP for Human Thromboplastin - rTF 16 : Data from UK NEQAS for Blood Coagulation

PB1121 Laboratory INRs Determined with Commercial Thromboplastins in >450 Centres Before and after the Establishment of the IRP for Human Thromboplastin - rTF 16 : Data from UK NEQAS for Blood Coagulation

Monitoring of treatment with vitamin K antagonists: recombinant thromboplastins are more sensitive to factor VII than tissue‐extract thromboplastins

Background Differences regarding sensitivity to factor VII have been suggested for recombinant human and tissue-extract thromboplastins used for International Normalized Ratio (INR) measurement, but the evidence is scarce. Differences in FVII sensitivity are clinically relevant, as they can affect INR stability during treatment with vitamin K antagonists (VKAs). Objectives To determine whether commercial thromboplastins react differently to changes in FVII. Methods We studied the effect of addition of FVII on the INR in plasma by using three tissue-extract (Neoplastin C1+, Hepato Quick, and Thromborel S) and three recombinant human (Recombiplastin 2G, Innovin, and CoaguChek XS) thromboplastins. Three different concentrations of purified human FVII (0.006, 0.012 and 0.062 μg mL-1 plasma), or buffer, were added to five certified pooled plasmas of patients using VKAs (INR of 1.5-3.5). Changes in FVII activity were measured with two bioassays (Neoplastin and Recombiplastin), and relative INR changes were compared between reagents. Results After addition of 0.062 μg mL-1 FVII, FVII activity in the pooled plasmas increased by approximately 20% (Neoplastin) or 32% (Recombiplastin) relative to the activity in pooled normal plasma. All thromboplastins showed dose-dependent INR decreases. The relative INR change in the pooled plasmas significantly differed between the six thromboplastins. No differences were observed among recombinant or tissue-extract thromboplastins. Pooled results indicated that the FVII-induced INR change was greater for recombinant than for tissue-extract thromboplastins. Conclusions Differences regarding FVII sensitivity exist between various thromboplastins used for VKA monitoring. Recombinant human thromboplastins are more sensitive to FVII than tissue-extract thromboplastins. Therefore, thromboplastin choice may affect FVII-mediated INR stability.

Clinical usefulness of international normalized ratio calibration of prothrombin time in patients with chronic liver disease

The international normalized ratio (INR) may not be directly applicable to patients with liver disease. We aimed to establish an alternative INR calibration system for patients with liver disease and to evaluate the effect of their use in chronic liver disease patients. Eighty-two patients with liver cirrhosis (LC) were included, and their prothrombin times (PTs) were measured by using 5 commercial thromboplastins. Each of the thromboplastins was also assigned an international sensitivity index (ISIliver) by the plasmas from LC patients. INRvka, INRliver, model for end-stage liver disease (MELD)vka, MELDliver, Child-Pugh (Child)vka, and Childliver scores were calculated. The coefficient of variance of INRvka was significantly larger than that of INRliver (P<0.01). The mean difference in INRvka between the thromboplastins was also significantly larger than that in INRliver (P<0.01). The total mean MELDliver score was higher than the total mean MELDvka score. The mean difference between the MELDvka and MELDliver scores (MELD score≥15) was 3.2%. We reconfirmed that the use of the alternative calibration system described herein for patients with liver disease may resolve the variability of INR measurement. Our data suggest that we would need to reevaluate the correlation between Child-Pugh class, MELD score, and clinical prognosis by using INRliver for patients with LC.

Usefulness and accuracy of the international normalized ratio and activity percent of prothrombin time in patients with liver disease

In order to determine the most reliable reporting style for prothrombin time (PT) in patients with liver disease, we examined the correlations between the plasma antigen levels of clotting factors and the PT activity percent or two international normalized ratios (INR), and compared the inter-reagent variation among these PT reporting styles. The PT was measured in 81 patients with liver diseases, including acute liver failure (ALF) (n = 10), acute liver injury (n = 52), chronic hepatitis (n = 8) and liver cirrhosis (n = 11), and in 75 warfarin-treated patients and 32 healthy volunteers. The PT of each plasma sample was determined with four commercial thromboplastins using an automated photo-optical coagulometer. For individual thromboplastin reagents, a locally determined international sensitivity index (local ISI) was derived using plasma obtained from healthy volunteers, warfarin-treated patients and liver disease patients. The INRW and INRLD were calculated using the corresponding local ISI. The PT activity percent was calibrated according to the Lineweaver-Burk equation. The PT values were compared with the plasma antigen levels of clotting factors X, II and VII measured using the enzyme-linked immunoassay method. The plasma factor X level was selected as the gold standard for measuring the synthetic liver function among the three clotting factors due to its significant relationship (P = 0.007) with the prognosis of ALF. The INRLD exhibited the closest correlation to the factor X level (r = 0.723-0.759), with the smallest inter-reagent variation among these reporting styles. The INRLD is the most appropriate PT reporting style for use in patients with liver disease.

Monitoring Coumadin-The Original Oral Anticoagulant

1. David L. McGlasson, MS, MLS(ASCP)CM[⇑][1] 1. 59th Clinical Research Division Laboratory Services, Wilford Hall Ambulatory Service Center, Lackland AFB, TX 1. Address for Correspondence- David L. McGlasson MS, MLS(ASCP)CM, Clinical Research Scientist, 59th Clinical Research Division Laboratory Services, Wilford Hall Ambulatory Service Center, Lackland AFB, TX 78236-9908, 210-292-6555, David.mcglasson{at}us.af.mil 1. Discuss the use of the prothrombin time (PT)/international normalized ratio (INR) for monitoring subjects on the oral anticoagulant Coumadin. 2. Be aware of the relevance of clinical and genetic data (CYP2C9, VKORC1) in predicting Coumadin dosing in subjects requiring different INR ranges. 3. Be knowledgeable of the chromogenic factor X (CFX) assay for monitoring Coumadin in subjects with lupus anticoagulants, transitioning subjects to Coumadin from direct thrombin inhibitors and for accurately following patients with elevated INRs. The prothrombin time (PT) has been the primary screening test for the laboratory evaluation of patients with an acquired or inherited coagulation factor deficiency of what was originally known as the extrinsic or common pathway of coagulation. It is used in conjunction with the international normalized ratio (INR) to monitor oral anticoagulant therapy (OAT) subjects on the oral anticoagulant, Coumadin.1 Tissue factor (TF) in vivo activates the coagulation cascade through the formation of the TF/FVIIa complex. The PT adds a TF/calcium mixture to citrated platelet-poor-plasma (PPP) in vitro to obtain the time for clot formation. There are many variables such as proper specimen collection, multiple reagent/instrument combinations and, most importantly, the sources of thromboplastin, usually rabbit and recombinant tissue factor, selected for the assay.1,2 The INR and Local INR Calibration The commercial thromboplastins vary widely in their sensitivities to Coumadin. Instrumentation differences (photo-optic or mechanical clot detection) may also play a clinically significant role in the performance of the PT/INR assay. With so many variables, providers treating subjects on OAT can be confused when comparing patient results from a number of laboratories using different reagent/instrument combinations. A facility that uses a high sensitivity thromboplastin will generate longer PT results than an assay that uses a low sensitivity thromboplastin. Therefore a subject on OAT may have a PT of 14 seconds with a low sensitivity reagent or 18 seconds with a more sensitive thromboplastin. Hence a subject monitored with insensitive thromboplastins would appear to require a higher dosage of Coumadin to… ABBREVIATIONS: CFX - chromogenic factor X; CLSI - Clinical and Laboratory Standards Institute; CYP2C9 - cytochrome P450; DTI - direct thrombin inhibitor; FII - coagulation factor II (prothrombin); FX - coagulation factor X; GMNPT - geometric mean normal prothrombin time; INR - international normalized ratio; IRP - international reference preparation; ISI - international sensitivity index; LA - lupus anticoagulant; OAT - oral anticoagulation therapy; PGX - pharmacogenetics; PST - patient self-testing; PT - prothrombin time; TTR - time in therapeutic range; VKORC1 - vitamin K epoxide reductase complex subunit 1; WHO - World Health Organization 1. Discuss the use of the prothrombin time (PT)/international normalized ratio (INR) for monitoring subjects on the oral anticoagulant Coumadin. 2. Be aware of the relevance of clinical and genetic data (CYP2C9, VKORC1) in predicting Coumadin dosing in subjects requiring different INR ranges. 3. Be knowledgeable of the chromogenic factor X (CFX) assay for monitoring Coumadin in subjects with lupus anticoagulants, transitioning subjects to Coumadin from direct thrombin inhibitors and for accurately following patients with elevated INRs. [1]: #corresp-1

The prothrombin time/international normalized ratio (PT/INR) Line: derivation of local INR with commercial thromboplastins and coagulometers – two independent studies

The WHO scheme for prothrombin time (PT) standardization has been limited in application, because of its difficulties in implementation, particularly the need for mandatory manual PT testing and for local provision of thromboplastin international reference preparations (IRP). The value of a new simpler procedure to derive international normalized ratio (INR), the PT/INR Line, based on only five European Concerted Action on Anticoagulation (ECAA) calibrant plasmas certified by experienced centres has been assessed in two independent exercises using a range of commercial thromboplastins and coagulometers. INRs were compared with manual certified values with thromboplastin IRP from expert centres and in the second study also with INRs from local ISI calibrations. In the first study with the PT/INR Line, 8.7% deviation from certified INRs was reduced to 1.1% with human reagents, and from 7.0% to 2.6% with rabbit reagents. In the second study, deviation was reduced from 11.2% to 0.4% with human reagents by both local ISI calibration and the PT/INR Line. With rabbit reagents, 10.4% deviation was reduced to 1.1% with both procedures; 4.9% deviation was reduced to 0.5% with bovine/combined reagents with local ISI calibrations and to 2.9% with the PT/INR Line. Mean INR dispersion was reduced with all thromboplastins and automated systems using the PT/INR Line. The procedure using the PT/INR Line provides reliable INR derivation without the need for WHO ISI calibration across the range of locally used commercial thromboplastins and automated PT systems included in two independent international studies.

The validity of the INR system for patients with liver disease

I read with interest the paper by Wei et al. [1], published recently in the Journal of Thrombosis and Thrombolysis on the assessment of the validity of the INR system for patients with liver disease and wish to make comments. The authors measured the INR with different commercial thromboplastins for a cohort of patients with liver disease and a cohort of patients stabilized on vitamin K antagonists (VKA). While the between-thromboplastin INRs for the patients with liver disease were significantly different, those for the patients on VKA were not. The obvious conclusion is that the INR (as it is calibrated for patients on VKA and here called INRvka) is not valid for patients with liver disease and, therefore, the model of end stage liver disease (MELD) score, once proposed as an objective index to prioritize patients for liver transplantation, would not allow parity of organ allocation. In their discussion Wei et al. [1], ignored at least seven of the many papers published over the last few years on this topic. Here few examples. Trotter et al. [2], were among the first in 2004 to show that the MELD score was not effective in securing parity of organ allocation. Other papers have consistently shown (from 1994 to 1999) that the INR, devised for patients on VKA, is not valid for patients with chronic liver disease [3, 4]. In 2007, Tripodi et al. [5] and Bellest et al. [6] reported independently on the same issue of Hepatology that the INRvka is not valid for patients with chronic liver disease as shown by the fact that the MELD, calculated by including in the equation the INRvka, depends on the thromboplastin used for testing. In the same papers these authors have independently shown that an alternative system of ISI calibration, provisionally called ISIliver (as opposed to the ISIvka) [5] can be obtained by inserting into the calibration plot, plasmas from patients with chronic liver disease instead of plasmas from patients on VKA [5, 6]. This alternative system of calibration proved effective in minimizing between-thromboplastin MELD results. More recently, Sermon et al. [7], confirmed these results, and Tripodi et al. [8] extended this model of ISIliver calibration to portable coagulation monitors. Finally, a review article on this topic has been published in the Journal of Thrombosis and Haemostasis in 2009 [9] and official recommendations have been issued independently by the International Society on Thrombosis and Haemostasis [10] and by the American Journal of Transplantation [11]. Surprisingly, some of the above papers [2–6, 8, 9] that appeared in the literature well before the publication of Wei et al. [1] have escaped their attention.

Results of the performance verification of the CoaguChek XS system

Background This is the first paper reporting a performance verification study of a point-of-care (POC) monitor for prothrombin time (PT) testing according to the requirements given in chapter 8 of the International Organization for Standardization (ISO) 17593:2007 standard “ Clinical laboratory testing and in vitro medical devices — Requirements for in vitro monitoring systems for self-testing of oral anticoagulant therapy”. The monitor under investigation was the new CoaguChek XS system which is designed for use in patient self testing. Its detection principle is based on the amperometric measurement of the thrombin activity generated by starting the coagulation cascade using a recombinant human thromboplastin. Methods The system performance verification study was performed at four study centers using venous and capillary blood samples on two test strip lots. Laboratory testing was performed from corresponding frozen plasma samples with six commercial thromboplastins. Samples from 73 normal donors and 297 patients on oral anticoagulation therapy were collected. Results were assessed using a refined data set of 260 subjects according to the ISO 17593:2007 standard. Results Each of the two test strip lots met the acceptance criteria of ISO 17593:2007 versus all thromboplastins (bias − 0.19 to 0.18 INR; > 97% of data within accuracy limits). The coefficient of variation for imprecision of the PT determinations in INR ranged from 2.0% to 3.2% in venous, and from 2.9% to 4.0% in capillary blood testing. Capillary versus venous INR data showed agreement of results with regression lines equal to the line of identity. Conclusion The new system demonstrated a high level of trueness and accuracy, and low imprecision in INR testing. It can be concluded that the CoaguChek XS system complies with the requirements in chapter 8 of the ISO standard 17593:2007.

Thromboplastin Composition Affects the Sensitivity of Prothrombin Time (PT) Clotting Tests To Direct Factor Xa Inhibitors.

Introduction: Thromboplastin reagents used for prothrombin time (PT) clotting assays vary in their sensitivity to anticoagulant drugs that directly inhibit Factor Xa (FXa). The International Sensitivity Index (ISI)/International Normalized Ratio (INR) system was introduced for monitoring warfarin, and corrects for differences in PT assay sensitivity. However, it does not adequately correct for differences in assay sensitivity to direct FXa inhibitors. The objective of this study was to determine how the composition of thromboplastin reagents affects PT sensitivity to the novel oral, direct FXa inhibitor rivaroxaban and how this correlates with the INR.Methods: Several recombinant thromboplastin reagents were prepared using different concentrations of NaCl, tissue factor and phospholipids (PL). They also contained different % of phosphatidylserine (PS), phosphatidylethanolamine (PE) and phosphatidylcholine (PC). These locally prepared thromboplastin reagents and five commercial thromboplastin assays were evaluated. PT ratios (PTR = PT with drug/PT without drug) were measured using normal human plasma to which rivaroxaban 1 μg/mL was added in vitro. Some PTRs were converted to INRs using locally determined ISI.Results: PT obtained with commercial thromboplastins was prolonged by rivaroxaban (Table), but the magnitude varied more than 3-fold, depending on the thromboplastin. Converting PTR to INR failed to normalize these results and made discrepancies more pronounced. Using locally prepared thromboplastin reagents, the PT sensitivity toward rivaroxaban was found to increase by decreasing the concentration of tissue factor or by increasing the concentration of PL or NaCl. Increasing the % PS generally decreased rivaroxaban sensitivity, while including PE generally increased rivaroxaban sensitivity. There was also a trend toward higher rivaroxaban sensitivity as the baseline PT increased. As with commercial thromboplastins, converting these PTR values to INR frequently made the discrepancies more pronounced.Conclusions: Changing the composition of thromboplastin reagents had disparate effects on the sensitivity of PT clotting tests to rivaroxaban. Furthermore, converting PTR values to INR failed to eliminate, and in some cases even exacerbated, the apparent differences in assay sensitivity of these PT clotting tests to rivaroxaban. This study sheds new light on the shortfall of the ISI/INR system to adequately correct for variation in the sensitivity of PT tests to direct FXa inhibitors. However, data show that other global clotting tests, such as PT, can be used to assess the efficacy of direct FXa inhibitors.Table 1:PT results for normal plasma spiked with 1 μg/ml rivaroxaban using commercial thromboplastinsCommercial thromboplastinsISIPTRINRRecombiplastin0.945.074.60Innovin0.982.252.22Thromborel S1.072.372.52Neoplastine CL+1.097.328.76Thromboplastin C+1.503.777.31INR, International Normalized Ratio; ISI, International Sensitivity Index; PTR, prothrombin time ratio

Comparison of local International Sensitivity Index calibration and 'Direct INR' methods in correction of locally reported International Normalized Ratios: an international study.

It is no longer feasible to check local International Normalized Ratios (INR) by the World Health Organization International Sensitivity Index (ISI) calibrations because the necessary manual prothrombin time technique required has generally been discarded. An international collaborative study at 77 centers has compared local INR correction using the two alternative methods recommended in the Scientific and Standardization Committee of the International Society on Thrombosis and Haemostasis guidelines: local ISI calibration and 'Direct INR'. Success of INR correction by local ISI calibration and with Direct INR was assessed with a set of 27 certified lyophilized plasmas (20 from patients on warfarin and seven from normals). At 49 centers using human thromboplastins, 3.0% initial average local INR deviation from certified INR was reduced by local ISI calibration to 0.7%, and at 25 centers using rabbit reagents, from 15.9% to 7.5%. With a minority of commercial thromboplastins, mainly 'combined' rabbit reagents, INR correction was not achieved by local ISI calibration. However, when rabbit combined reagents were excluded the overall mean INR deviation after correction was reduced further to 3.9%. In contrast, with Direct INR, mean deviation using human thromboplastins increased from 3.0% to 6.6%, but there was some reduction with rabbit reagents from 15.9% to 10% (12.3% with combined reagents excluded). Local ISI calibration gave INR correction for the majority of PT systems but failed at the small number using combined rabbit reagents suggesting a need for a combined reference thromboplastin. Direct INR correction was disappointing but better than local ISI calibration with combined rabbit reagents. Interlaboratory variability was improved by both procedures with human reagents only.

Reliability of the international normalized ratio for monitoring the induction phase of warfarin: Comparison with the prothrombin time ratio

The International Normalized Ratio (INR) was introduced to reduce the variability of prothrombin time (PT) reporting. One potential problem with the use of the INR is the assumption that its reliability is reduced when it is used to monitor patients during the induction phase of treatment. This shortcoming arises because the model used to establish the INR system is based on the use of pooled plasma from patients stabilized on warfarin for at least 6 weeks. Because the prolongation of the PT by warfarin during the induction phase mainly reflects reduction in factor VII levels (whereas the prolongation of the PT after 6 weeks of stabilization reflects reductions in factors X, II, and VII), there exists a potential for loss of accuracy of the INR during warfarin induction. To overcome this potential problem, it has been suggested that the PT ratio should be used to report results during the induction phase of treatment and that the INR system should be reserved for reporting results after the patient has been stabilized. This approach is confusing to the clinician. In addition, the validity of this approach has never been demonstrated in a clinical study. To address this issue, we studied 43 patients for the first 5 days after they started warfarin therapy. We measured the PT in the same plasma samples from each patient with five different commercial thromboplastins. The variance in the PT ratios among the five thromboplastins was compared with the variance obtained with the INR values derived from the PT ratios when using the international sensitivity indexes provided by the manufacturer. Our results indicate that, even during the induction phase, there is less variance with the INR system than with the PT ratio system.

Prothrombin Time Sensitivity and Specificity to Mild Clotting Factor Deficiencies of the Extrinsic Pathway: Evaluation of Eight Commercial Thromboplastins

Prothrombin-time (PT) sensitivity and specificity to mild clotting factor II, V, VII and X deficiencies have rarely been studied. We therefore carried out a prospective study, in 350 patients, of eight commercial thromboplastins (CTs) in their ability to detect mild clotting factor deficiencies, notably in factor VII. In each patient the factor II, V, VII and X clotting activities and PT performed with each CT were determined. For each CT, PT sensitivity and specificity in detecting factor deficiencies below 0.5 U/ml or below 0.4 U/ml were determined at various PTs, and then Receiver Operator Characteristic curves constructed. At optimum PT threshold level (sensitivity = specificity), exactitude varied from 0.64 to 0.74 (p < 0.01) and from 0.67 to 0.81 (p < 0.0001) in detecting deficiencies below 0.5 and 0.4 U/ml respectively. In conclusion, this study shows the limits of the PT test as performed with 8 CTs in patients with mild clotting factor deficiencies. The impact of such differences in sensitivity and specificity on monitoring certain patients subjects to decrease in coagulation factor, and, in particular, of those under low dose oral anticoagulant, remains to be determined.

Effect of thromboplastin and coagulometer interaction on the precision of the International Normalised Ratio.

To examine the magnitude of thromboplastin and coagulometer interactions on the precision of International Normalised Ratio (INR) values when the manufacturers' recommended instrument specific International Sensitivity Index (ISI) values are adopted for the INR calculation. The variability of INR values obtained from four automated phototopical coagulometers frequently used in North American laboratories was studied. When used with five commercial thromboplastins of moderate to high sensitivity (ISI values 0.92-1.97), 20 prothrombin time results were generated for each of a population of 98 patients on established warfarin treatment. The mean INR values of the patients ranged from 2.05 to 2.81, depending on which reagent/coagulometer combination was used. Within patient variation increased as the median INR value increased. The mean coefficient of variation of within patient INR values was 10%; the mean coefficient of variation of the prothrombin time results in seconds and prothrombin time ratio were 21 and 18%, respectively. There was considerable bias in the estimated ISI values of the thromboplastins compared with the manufacturers' instrument specific ISI value. Despite this apparent imperfection, our study clearly showed that the INR is preferable to other prothrombin time reporting formats for assessing the degree of anticoagulation for patients on warfarin treatment.

Effect of Concentration of Trisodium Citrate Anticoagulant on Calculation of the International Normalised Ratio and the International Sensitivity Index of Thromboplastin

The aim of this study was to determine whether the concentration of trisodium citrate used to anticoagulate blood has an effect on the INR of the sample and the ISI of the thromboplastin. Five thromboplastins including and Australian reference material were used to measure the prothrombin time of normal and patient samples collected into two concentrations of trisodium citrate--109 mM and 129 mM. There was no effect of citrate concentration on the INRs determined with the reference material. However for the other four thromboplastins there was a significant difference between INRs for the two citrate groups. The prothrombin times of the samples collected into 129 mM were longer than those collected into 109 mM. This difference was only slight in normal plasma but more marked in patients receiving oral anticoagulants, causing the INRs for patient plasmas collected into 129 mM citrate to be higher then the corresponding samples collected into 109 mM citrate. From orthogonal regression of log prothrombin times by the reference method against each thromboplastin, we found that the ISI for each thromboplastin was approximately 10% lower when determined with samples collected into 129 mM citrate than with samples collected into 109 mM. These results suggest that the concentration of trisodium citrate used for collection of blood samples can affect the calculation of the INR and the calibration of the ISI of thromboplastin. This was found both for commercial thromboplastins prepared by tissue extraction and for a recombinant tissue factor.

Oral Anticoagulants: Mechanism of Action, Clinical Effectiveness, and Optimal Therapeutic Range

The optimal therapeutic range for oral anticoagulant therapy was reviewed by the Committee on Antithrombotic Therapy of the American College of Chest Physicians (ACCP) and the National Heart, Lung and Blood Institute (NHLBI) in 1986 and again in 1989. 1 ACCP/NHLBI National conference on antithrombotic therapy. Chest 1989; 89(suppl): 1-169 Google Scholar At that time, a recommendation was made that the intensity of warfarin treatment should be reduced for many indications. 1 ACCP/NHLBI National conference on antithrombotic therapy. Chest 1989; 89(suppl): 1-169 Google Scholar Since then, new clinical trials have been published which support these recommendations, and the optimal therapeutic range for many indications has been clarified. Nevertheless, the control of oral anticoagulant therapy remains a subject of confusion. The confusion persists for two related reasons. The first is the marked variation in the responsiveness of different commercial thromboplastin reagents to reductions by warfarin of vitamin K dependent clotting factors 2 Zucker S Cathey MH Sox PJ et al. Standardisation of laboratory tests for controlling anticoagulant therapy. Am J Clin Pathol. 1970; 53: 348-354 Crossref PubMed Scopus (30) Google Scholar , 3 Poller L Progress in standardisation in anticoagulant control. Hematol Rev. 1987; 1: 225-241 Google Scholar , 4 Bailey EL Harper TA Pinkerton PH Therapeutic range of one-stage prothrombin time in control of anticoagulant therapy. Can Med Assoc J. 1971; 105: 1041-1043 PubMed Google Scholar , 5 Latallo ZS Thomson JM Poller L An evaluation of chromogenic substrates in the control of oral anticoagulant therapy. Br J Haematol. 1981; 47: 307-318 Crossref PubMed Scopus (21) Google Scholar , 6 Poller L Taberner DA Dosage and control of oral anticoagulants: an international survey. Br J Haematol. 1982; 51: 479-485 Crossref PubMed Google Scholar , 7 Verstraete M Clark PA Wright IS Use of different tissue thromboplastins in the control of anticoagulant therapy. Circulation. 1957; 16: 213-226 Crossref PubMed Scopus (6) Google Scholar measured in the prothrombin time (PT) test; and the second is the reluctance of hematologists in North America to adopt the standardized international normalized ratio (INR) method of reporting which is now accepted internationally. In this update on oral anticoagulant therapy, we have made a number of changes. The first is to report our recommendations for therapeutic ranges of INR since recent reports indicate that the variations in responsiveness of commercial thromboplastins is so wide that the term typical North American thromboplastin is no longer valid. The second is to recommend an INR of 2.0 to 3.0 for all indications (including recurrent systemic embolism) except mechanical prosthetic heart valves for which an INR of 2.5 to 3.5 is recommended. The intensity of the therapeutic range has been lowered for mechanical prosthetic valves because of reports that a low intensity regimen is effective for this indication and because the risk of bleeding increases with increasing anticoagulant intensity. The third change is an expansion of this chapter to include mechanism of action of oral anticoagulants, their pharmacokinetics, practical dosing considerations, and the management of patients who require reversal or lowering of anticoagulant intensity.

The Dependence of the International Sensitivity Index on the Coagulometer Used to Perform the Prothrombin Time

This study was designed to detect any effect that different types of coagulation instrument may have on the International Sensitivity Index (ISI) of a thromboplastin. Manufacturers of commercial thromboplastins now calibrate their reagents against the World Health Organization international reference preparation to assign them an ISI. This enables the prothrombin time (PT) estimated with that reagent to be expressed as an International Normalised Ratio (INR). One batch of Thromborel S was calibrated against the Australasian Reference Thromboplastin (ART). The Thromborel S was used on three photo-optical instruments, the Automated Coagulation Laboratory (ACL) (Instrumentation Laboratory), the Cobas Fibro (Roche), and the Coag-a-Pet (General Diagnostics). PTs using ART were performed manually using the reference method. The ISIs calibrated in our laboratory when the ACL and Cobas Fibro were used were not significantly different at the 95% level, being 1.102 +/- 0.018 and 1.134 +/- 0.022 respectively. The ISI with the Coag-a-Pet of 1.223 +/- 0.023 was significantly different to that of the ACL and the Cobas Fibro at the 95% level. The flowcharts for a computer program to perform the necessary calculations are provided. The program allows for the entry and editing of data from the calibration procedure, and provides a mean normal PT and normal range, the ISI and 95% confidence limits of the calibration, and a chart for the conversion of the test PTs to INRs. The authors have made available an IBM compatible program for the calibration of thromboplastins.

Long-term stability of international reference preparations for thromboplastins.

Three certified reference materials for thromboplastins are available from the Community Bureau of Reference (BCR) of the European Commission for calibration of commercial thromboplastins used for control of oral anticoagulant therapy. The long-term stability of these reference materials has been monitored by two independent laboratories, using deep-frozen and lyophilized plasma samples. Prothrombin times and prothrombin time ratios measured on 19 occasions in the period 1981-86 have been analysed for trend with time. Although significant trends of prothrombin time and ratio (P less than 0.05) were observed, a consistent pattern of trends could not be recognized. The significant trends of prothrombin time and prothrombin time ratio are most probably due to changes in local laboratory conditions. There is no indication that the reference materials have deteriorated since the beginning of the study. It is recommended that long-term stability monitoring of thromboplastins be performed by at least two laboratories simultaneously.

Thrombomodulin activity in commercial thromboplastin preparations

Recently, it was reported that endothelial cells contain a membrane protein which serves as a cofactor for the activation of protein C by thrombin (thrombomodulin). Because many commercial thromboplastins are prepared from vessel-rich tissues/organs (lung, brain, placenta), it was hypothesized that some preparations may contain significant amounts of thrombomodulin; theoretically, thrombomodulin contaminations may enhance the prothrombin time. Among 8 different commercial thromboplastins tested two thromhoplastins were found to contain significant amounts of thrombomodulin activity: Simplastin (64 U/ml) and Ortho (12.8 U/ml). However, we were unable to demonstrate that the presence of the thrombomodulin activity in these thromboplastins could affect the prothrombin time either by delaying fibrin formation (formation of thrombin-thrombomodulin complexes) or by inhibiting activated factor V (formation of activated protein C).

The calibration of rabbit tissue thromboplastins: experience of the Dutch Reference Laboratory for anticoagulant control

A number of commercial rabbit tissue thromboplastins used in oral anticoagulant control have been calibrated against the first International Reference Preparation for thromboplastins. This was done in a three-stage procedure by one laboratory, each stage representing a different level of thromboplastin comparability. The calibration model recently recommended by ICTH and ICSH was tested. This model proved to be suitable, although a statistically significant aberration was observed for some of the thromboplastins. The bias introduced by using the model in these non-ideal cases was small compared to the overall variation of the International Normalized Ratio, being the universal scale for reporting the prothrombin time during oral anticoagulant control. Batch-to-batch calibration using lyophilized pooled plasmas could be reliably performed for several commercial thromboplastins.

Comparison of the spectrophotometric determination and the two-stage coagulation assay of tissue factor activity

The tissue factor activity of human brain thromboplastin and 6 commercial thromboplastins was determined by a spectrophotometric method and a two-stage coagulation assay. The thromboplastins were incubated with an excess of Factor VII and Factor X, and the activation of Factor X was estimated from the rate of hydrolysis of the chromogenic substrate S-2222 and from the coagulation time of plasma enriched in phospholipids. The results obtained by the two methods were related linearly and showed a correlation coefficient of 0.89. The coefficient of variation was 11% in the spectrophotometric method and 25% in the two-stage coagulation assay.