European Concerted Action on Anticoagulation (ECAA). An assessment of lyophilised plasmas for ISI calibration of CoaguChek and TAS whole blood prothrombin time monitors

AIMS: The recommended method for the international sensitivity index (ISI) calibration of whole blood point of care testing (POCT) prothrombin time (PT) systems was originally described by Tripodi et al in 1993 but is too complex and demanding. The present European Concerted Action on Anticoagulation (ECAA) study aimed to assess the reliability of simpler ISI calibration using lyophilised plasma samples. METHODS: ISI calibrations using three different types of ECAA lyophilised plasma samples (artificially depleted, individual, and pooled coumarin) were compared with whole blood calibrations on CoaguChek Mini and TAS PT-NC POCT monitors at 10 centres. RESULTS: With CoaguChek Mini systems, lyophilised coumarin plasma samples (both single donation and pooled) gave ISI and international normalised ratio (INR) values comparable to whole blood. With artificially depleted plasma, ISI and INR values were too high. With TAS PT-NC systems, all three types of lyophilised plasma samples gave inaccurate ISI and unreliable INR results, similar to previous ECAA findings with fresh plasma calibrations. CONCLUSIONS: With CoaguChek Mini systems, ISI calibration can be simplified by the use of ECAA lyophilised plasma samples from coumarin treated patients. Further study is needed to devise a simpler calibration method for the TAS PT-NC system.     

European concerted action on anticoagulation. Minimum numbers of lyophilized plasma samples for ISI calibration of CoaguChek and TAS point-of-care whole blood prothrombin time monitors

International sensitivity index (ISI) calibration of whole blood prothrombin time (PT) monitors is too complex. We previously simplified the method by using European Concerted Action on Anticoagulation (ECAA) lyophilized plasma samples with the TAS PT-NC (Bayer AG, Leverkusen, Germany) and the CoaguChek Mini (Roche Diagnostics, Mannheim, Germany) whole blood PT monitoring systems. The TAS PT-NC required a correction derived from the line of equivalence. Monte Carlo bootstrap analysis of reducing numbers of test samples was performed with both systems. Plasma samples from patients receiving coumarin (coumarin samples), healthy subjects (normal samples), and plasma samples artificially depleted of coagulation factors were used. With the TAS PT-NC, 20 coumarin samples or 20 artificially depleted samples with 7 normal samples gave reliable ISI and international normalized ratio and satisfactory precision. With the CoaguChek Mini, 30 coumarin and 10 normal samples were required. Simplification of ISI calibration of the 2 monitoring systems is possible using fewer ECAA lyophilized plasma samples than the 80 required according to the World Health Organization guidelines for conventional PT systems and previously recommended for fresh plasma samples tested on the same 2 monitoring systems.     

The importance of "like to like" ISI calibrations with freeze dried plasmas. European Concerted Action on Anticoagulation.

AIM: To assess reliability of like to like and cross species calibrations using two types of certified freeze dried plasma calibrants--artificially depleted of vitamin K clotting factors, and from coumarin treated patients. METHODS: Six ECAA national control laboratories provided certified values for the freeze dried plasmas in terms of the human plain international reference preparation (IRP) (BCT/441) with the manual prothrombin time technique. Eight other ECAA national laboratories determined international sensitivity index (ISI) values in full fresh plasma same species and cross species WHO calibrations against a low ISI human IRP (BCT/441) of the ECAA low ISI human thromboplastin and high ISI ECAA rabbit thromboplastin. Parallel calibrations were performed using the certified values. RESULTS: Calibrations on fresh plasmas of the human ECAA reference thromboplastin (stated ISI = 0.95) gave ISI of 0.957 against the human IRP and 1.66 against the rabbit IRP. The ECAA rabbit (stated ISI = 1.67) gave an identical value on the fresh plasma calibration v the human IRP. With freeze dried depleted plasmas certified in terms of the human IRP, the ISI of the ECAA human was 1.01, but the ECAA rabbit (stated ISI = 1.67) gave a low ISI of 1.47. The freeze dried coumarin plasmas gave an ISI of 0.943 for the ECAA human but only 1.493 for the ECAA rabbit. CONCLUSIONS: Fresh plasmas give reliable ISI when calibrating thromboplastins in same species and cross species calibrations. Freeze dried plasmas certified in terms of a single IRP, whether artificially depleted or of coumarin plasma origin, cannot be used for calibration of dissimilar thromboplastins.     

International sensitivity index calibration of the near-patient testing prothrombin time monitor,ProTime

Near-patient testing devices (monitors) capable of measuring prothrombin time on an unmeasured drop of blood would be suitable alternatives to centralized laboratory monitoring of patients receiving oral anticoagulants. The essential prerequisite for the use of these monitors is their conformity to the international sensitivity index (ISI) calibration model recommended by the World Health Organization. We report on the ISI calibration of the ProTime monitor (International Technidyne, Edison, NJ) designed and approved for patient self-testing. According to our results, this monitor can be calibrated by adopting the model already used for other monitors. The apparent ISI was close to unity. Overall, the international normalized ratio values displayed by the monitor agreed with those measured with the international reference preparation for thromboplastin. Confirmation of these results in a large multicenter study is warranted.     

Suggestions and propositions to resolve some issues for standardization of prothrombin time and activated partial thromboplastin time]

Prothrombin time (PT) and activated partial thromboplastin time (APTT), popularized as a routine assay for screening blood coagulation disorders and monitoring anticoagulant therapy, still involve some issues regarding standardization. In this lecture, we present propositions to resolve these problems in respective laboratory. Although international normalized ratio (INR) calculated by international sensitivity index (ISI) of PT reagent seems to improve discrepancy of sensitivity between reagents, local calibration of sensitivity of PT reagent in respective laboratories (local SI) is reasonable to make INR/ISI system more useful. However, local calibration of reagent is not easy by WHO recommended method in a small size laboratory. By using AK calibrant (IMMUNO AG), one of calibration plasma for INR, we investigated its possibility to calibrate local SI in four different reagents, compared with the recommended methodology. The results led the following process to determine reagent and calibrate local SI for practical use of INR/ISI system. (a) Use PT reagent of which ISI is close to 1.0 if possible, and utilize manufacture's ISI as is for INR. (b) Select PT reagent labeled specific ISI for an instrument as the same as used in the lab., and use the manufacture's ISI as is, if impossible to choose small ISI reagent. (c) If use a reagent of which ISI is close to 2.0 and shown no specific ISI for used detector, adjustment of local SI by commercial calibration plasma is recommended when unavailable warfarinized patient plasma. In APTT, we attempted to evaluate sensitivity between five different APTT reagents with a patient model by hemophilia A plasma contained various FVIII: C. This model reflected difference of sensitivity between reagents in results. Because standardization of APTT is not improved in this point, certification of APTT pattern in each laboratories with patient models is required for not only monitoring of heparinization, but also screening of typical coagulation disorders such as hemophilia and von Willebrand disease.     

Influence of blood collection systems on the prothrombin time and international sensitivity index determined with human and rabbit thromboplastin reagents

Three brands of blood collection tubes were studied for their influence on the prothrombin time (PT) and international sensitivity index (ISI) for 5 commercial thromboplastin reagents. With all reagents, PTs were shorter in Vacutainer (Becton Dickinson Vacutainer Systems, Plymouth, England) samples than in S-Monovette (Sarstedt, Nümbrecht, Germany) or Venosafe (Terumo Europe, Leuven, Belgium) samples. ISI values were higher with Vacutainer samples than with S-Monovette or Venosafe samples. The ISI differences between the tubes were small for Thromborel-S (2.1%; Dade Behring, Marburg, Germany) and Hepato Quick (1.1%; Diagnostica Stago, Asnières, France; Roche Diagnostics Nederland, Almere, the Netherlands) but greater for Neoplastin Plus (5.5%; Diagnostica Stago; Roche Diagnostics Nederland), Simplastin HTF (8.3%; bioMérieux, Durham, NC), and Innovin (8.8%; Dade Behring). The PT and ISI differences between the tubes could be explained mostly by the effect of magnesium ion contamination in the sodium citrate solutions. When PT ratios were transformed into international normalized ratios (INRs) using crossover ISI (ie, samples collected with one type of tube and ISI determined with another collection system for the PT reagent), the differences in mean INRs could be approximately 10%. For ISI calibration of reference thromboplastins, blood collection tubes should be used with minimal divalent metal ion contamination of the citrate solution.     

Time to think outside the box? Prothrombin time,international normalised ratio,international sensitivity index,mean normal prothrombin time and measurement of uncertainty a novel approach to standardisation

BACKGROUND: The prothrombin time (PT) assay is the most clinically ordered coagulation test, and most often used for monitoring of vitamin K antagonist therapy (e.g., warfarin), where results are expressed as an international normalised ratio (INR). The INR is in essence the patient's PT 'mathematically adjusted' to a standardised value taking into account the peculiarities of the test system as defined by an ISI (international sensitivity index) and MNPT (mean normal prothrombin time). Although some manufacturers provide assigned ISI values for specific PT reagents and instrumentation, it is still recommended practice that laboratories check or validate these ISIs, as well as estimate the MNPT. Where an ISI is not provided by a manufacturer, the laboratory needs to estimate its own value. Current recommendations suggest the use of commercial reference-plasma calibration sets, but there is limited information on the performance of these in the field. RESULTS: We report a comparative study that assessed the utility of three such commercial calibration plasma sets, used as recommended, as well as alternate or supplementary procedures for estimation of ISI and MNPT. The latter included one novel approach using comparative data of 'existing' versus 'replacement' reagent, as well as assessment of external quality assurance data. Although MNPT value estimates were not grossly disparate, a wide variety of ISI values (e.g., 1.12-1.30 for our primary instrument) was obtained with the different plasma sets. CONCLUSION: Because of the above, further verification checks are required prior to acceptance of ISI and MNPT estimates generated from commercial plasma calibration sets. We also provide some recommendations regarding the process of standardisation of INR testing.     

The effects of freeze drying and freeze drying additives on the prothrombin time and the international sensitivity index

AIM: To determine whether freezing, freeze drying protective additives, or freeze drying of plasma samples from patients on coumarin treatment and from normal individuals affects prothrombin times or the international sensitivity index (ISI) calibration. METHODS: The effect of the addition of the protective additives singly and combined on the prothrombin time of coumarin samples and normal samples before and after freeze drying was observed using high and low ISI reference thromboplastins. ISI values were also determined. RESULTS: Freezing caused a prolongation of prothrombin time in the normal plasma samples with both reagents, which was significant with the low ISI human. Prolongation (non-significant) of the prothrombin time in coumarin plasma samples occurred with the human reagent only. Significant prolongation of normal prothrombin time by some of the protective additives before and after freeze drying was observed with both thromboplastins but to a greater extent with the human. Significant prolongation of prothrombin time in coumarin plasma samples was observed, but again was more marked with human thromboplastin. An approximate ISI was determined on the 20 coumarin samples. The only marked ISI change was with the WHO human thromboplastin after freeze drying of plasma, where a decrease from 0.95 to 0.90 was observed, corresponding to a marked prothrombin ratio increase. CONCLUSIONS: Freeze drying additives and the freeze drying procedure prolong normal and coumarin prothrombin times, with low ISI thromboplastin. Less marked prolongations occurred with a high ISI rabbit reagent, coumarin samples showing more significant prolongations. Marked ISI change in freeze dried plasma was only recorded with the low ISI ECAA human reagent. Frozen normal plasma samples cannot be used with confidence for ISI calibrations.     

Point-of-care testing for prothrombin time, but not activated partial thromboplastin time, correlates with laboratory methods in patients receiving aprotinin or epsilon-aminocaproic acid while undergoing cardiac surgery.

Point-of-care testing (POCT) of coagulation parameters can help optimize transfusion practice in cardiac surgery. Antifibrinolytic agents may interfere with the laboratory and/or POCT coagulation assays. This randomized controlled study compared coagulation parameters obtained from a whole blood POCT coagulation device with a typical laboratory instrument in cardiac surgery patients receiving aprotinin, epsilon-aminocaproic acid, or normal saline before undergoing cardiopulmonary bypass. Aliquots of arterial blood samples from 42 patients were collected perioperatively, and their prothrombin times (PTs) and activated partial thromboplastin times (aPTTs) were measured by POCT and laboratory instrumentation. Linear regression and error analyses were used for the method comparison. For PT, the POCT device compared favorably with the laboratory method. For aPTT, the POCT device did not compare well with the laboratory method. Treatment with antifibrinolytic agents does not interfere with determination of PT.     

Monitoring oral anticoagulant treatment with the TAS near-patient test system: comparison with conventional thromboplastins.

BACKGROUND: A number of instruments have been developed for determination of prothrombin time (PT) and International Normalised Ratio (INR) at locations not limited to central laboratories. AIM: To evaluate one such portable instrument, the Thrombolytic Assessment System (TAS), which can be used in a near-patient setting. METHODS: Samples from 20 normal subjects and 48 patients treated with warfarin for venous thromboembolic disease were studied. The warfarin group was divided into: initiation phase (n = 10), combined warfarin and heparin (n = 10), stabilised therapy (n = 20), and over anticoagulated patients (n = 8). PTs and INRs were determined in each group using three conventional thromboplastins (Diagen Activated, Manchester Reagent, and Instrumentation Laboratory) and two TAS techniques (whole blood or plasma). An independent International Sensitivity Index (ISI) calibration of the TAS system was performed. RESULTS: Calculated ISIs for the TAS were 1.028 and 0.984 for plasma and whole blood analysis, respectively, compared with manufacturer's values of 0.98 and 0.97. INR results with TAS (whole blood) were 11% less than those obtained with Diagen Activated (p < 0.01) and 16% less than those obtained with Instrumentation Laboratory (p < 0.001) when manufacturers' mean normal PT and ISI were used for TAS INRs. TAS (whole blood) results were similar to TAS plasma or Manchester Reagent results. The use of a locally determined mean normal prothrombin time (MNPT) improved agreement between TAS and the other reagents, abolishing the significant difference between INRs determined with TAS (whole blood) and Diagen Activated techniques. CONCLUSION: The TAS system can be used with whole blood or plasma and produces similar INRs to those obtained with Diagen Activated or Manchester Reagent using manufacturer's ISI and a locally determined MNPT. Results were lower with TAS or Manchester Reagent compared with those obtained with Instrumentation Laboratory thromboplastin.     

Automation and prothrombin time: a United Kingdom field study of two widely used coagulometers.

Current performance in the prothrombin time (PT) of the two main United Kingdom coagulometer/thromboplastin systems was assessed in a field survey. Twenty abnormal samples covering a wide spectrum of International Normalised Ratio (INR) were distributed to users of the KC4/KC10 and Coag-a-Mate instruments. Coagulometer results were compared with those of the manual method. A substantial minority with each system showed good agreement with the manual reference values. There was, however, a considerable variation between instruments, meaningful in clinical terms, evidenced by varying regression slopes and local system International Sensitivity Indices (ISI). For intense anticoagulation (3.0 to 4.5 INR) a larger dose of warfarin is needed with the Coag-a-Mate than with the KC instruments. With a manual INR of 4.0 the KC instruments tended to give longer PT (mean INR + 0.3); the Coag-a-Mate PT was generally shorter (mean INR -0.1). With both systems the mean normal PT were shorter than the manual but the degree of shortening did not parallel that of the abnormal samples. This effect undermines the use of a simple prothrombin ratio and of an INR value derived from it, based on a manual ISI. The use of a system related ISI cannot, however, be recommended until local instrument variables are controlled.     

Capillary whole blood testing by a new portable monitor. Comparison with standard determination of the international normalized ratio

We evaluated a new portable monitor (AvoSure PT PRO, Menarini Diagnostics, Firenze, Italy) developed to test the prothrombin time in capillary blood and plasma by comparing it with the standard laboratory determination. We studied 62 patients receiving acenocoumarol therapy. The international normalized ratio (INR) in capillary blood was analyzed by 2 methods: AvoSure PT PRO and Thrombotrack Nycomed Analyzer (Axis-Shield, Dundee, Scotland). Parallel studies were performed in plasma samples by a reference method using the Behring Coagulation Timer (Behring Diagnostics, Marburg, Germany). Plasma samples also were tested with the AvoSure PT PRO. Correlation was good for INR values for capillary blood and plasma samples by AvoSure PT PRO and our reference method (R2 = 0.8596) and for capillary blood samples tested by the AvoSure PT PRO and Thrombotrack Nycomed Analyzer (R2 = 0.8875). The correlation for INR in capillary blood and plasma samples by AvoSure PT PRO was 0.6939 (P < .0004). Capillary blood determinations are rapid and effective for monitoring oral anticoagulation therapy and have a high correlation to plasma determinations. AvoSure PT PRO is accurate for controlling INR in plasma and capillary blood samples, may be used in outpatient clinics, and has advantages over previous portable monitors.     

Screening INR deviation of local prothrombin time systems.

AIM: To assess the reliability of local international normalised ratios (INR) using a set of three international reference preparation (IRP) certified freeze dried plasmas. METHODS: 55 centres in the United Kingdom and the Republic of Ireland participated; 36 centres employed coagulometers and 19 a manual prothrombin time technique, all with the same batch of routine commercial thromboplastin. The plasmas had certified INR with the manual technique using a thromboplastin IRP, and results were provided graphically to participants for self assessment. An INR deviation of more than +/- 10% from the certified INR with any of the screening plasmas was regarded as unsatisfactory and clinically significant. Sets of 20 freeze dried plasmas were provided for local ISI calibrations and sets of seven freeze dried normals were provided for supplementary exercises where screening results were unsatisfactory. RESULTS: 15 of 38 coagulometers, but only three of the 19 manual prothrombin time test techniques, gave unsatisfactory results. With 10 of the 15 unsatisfactory coagulometer results the problem was resolved by local ISI calibrations with plasma calibrant sets provided. Unsatisfactory results with manual technique in all four instances were corrected by substitution of the mean result with freeze dried normal plasmas provided. CONCLUSIONS: The freeze dried plasma screening set was useful in detecting incorrect INR in a high proportion of coagulometer users and a smaller number of participants using the manual technique.     

Evaluation of European Concerted Action on Anticoagulation lyophilized plasmas for INR derivation using the PT/INR line

The prothrombin time/international normalized ratio (PT/INR) Line method based on 5 certified European Concerted Action on Anticoagulation (ECAA) plasmas provides reliable local INR values without conventional World Health Organization international sensitivity index calibrations. The present study investigated the use of different numbers and types of ECAA calibrant plasmas to derive accurate PT/INR Lines and reliable INR values. The numbers ranged from 3 to 10 plasmas in a set with normal or abnormal samples. Sets were selected, and sampling was repeated 1,000 times for each center to derive PT/INR Lines. The lines were selected randomly or from clusters. The INR values of 5 independent "validation" plasmas were compared before and after correction. In 56 calibrations, 5 ECAA plasmas gave better results than did fewer plasmas. Plasmas with wide-ranging INR values gave better results than randomly selected sets, and including a normal plasma was not essential. The INR deviations of validation plasmas from certified values were reduced with sets of human, bovine/combined, and rabbit reagents. Deviations of more than 10% from certified INR values were significantly reduced (P < .001).     

Multicentre evaluation of the Thrombotest International Sensitivity Index used with a steel ball coagulometer.

AIM: To compare the International Sensitivity Index (ISI) of the Thrombotest reagent used with a steel ball coagulometer (KC) to the ISI of the same reagent used with the manual (tilt tube) technique. METHODS: The study was carried out by eight laboratories using their own KC instrument and method of testing. All laboratories used the same batch of Thrombotest to determine the clotting times of fresh blood samples from 20 local healthy volunteers and 60 patients on long term oral anticoagulant therapy. KC clotting times were plotted against manual clotting times on double logarithmic scales. Orthogonal regression lines were calculated to assess the ISI. RESULTS: In two laboratories the ISI of the KC method was lower than that of the manual method; these differences, however, were 2% or less. In the other laboratories no clinically important differences were observed between ISI values obtained. However, the clotting times determined with the KC methods were shorter than the manual values. CONCLUSIONS: The ISI of Thrombotest determined with the KC methods was very similar to the manual value. Therefore, use of the ISI value supplied by the manufacturer without adjustment is justified. The mean normal prothrombin time, however, must be determined locally.     

A novel whole blood capillary technic for measuring the prothrombin time

The prothrombin time (PT) is frequently performed to monitor anticoagulant therapy. Although relatively simple to perform, it requires venipuncture and laboratory resources for sample handling and analysis. A recently developed capillary whole blood device that uses fingerstick samples was evaluated. Paired capillary whole blood and reference plasma PTs were performed in 858 samples from 732 subjects. The PT for normal volunteers (n = 193) was 11.8 +/- 0.9 seconds with the use of the new instrument and 12.1 +/- 0.5 seconds with the use of the reference method. In samples from 539 patients receiving anticoagulants, the correlation coefficient between the two methods was 0.96. Venous whole blood without anticoagulant and capillary whole blood gave equivalent results, which suggests that the fingersticks do not effect the quality of the specimen. Variation in hematocrit between 23.4% (0.34) and 53.8% (0.538) did not alter the performance of the instrument. The new instrument is easy to use and may allow testing by nonlaboratory personnel and patients. It obviates the need for venipuncture, provides immediate results, and appears to be comparable in accuracy to current reference methods.     

Effect of protein on hemoglobin and hematocrit assays with a conductivity-based point-of-care testing device: comparison with optical methods

Point-of-care testing (POCT) for blood hemoglobin and hematocrit (H/H) levels provides rapid patient assessment including the need for transfusion. Conductivity-based methods of blood H/H determinations can be influenced by plasma protein concentration. To assess this factor, we measured H/H levels at varying protein concentrations using two POCT instruments: iSTAT-1 (conductivity method) and Hemocue (optical method). These H/H results were compared to results obtained by our core laboratory hematology analyzer (GenS). Anticoagulated whole blood was centrifuged to sediment the red blood cells; the plasma was removed to serve as source of protein for mixing studies. A series of reconstituted samples was prepared with varying H/H and protein levels. To mimic hemodilution in clinical practice, samples were diluted with saline or lactated Ringer's solutions. Following H/H analysis, the samples were centrifuged and protein determined in the supernatant plasmas. The H/H results obtained with the Hemocue instrument correlated exactly with those of the GenS analyzer at protein concentrations from 0.7 to 6.2 g/dl. The correlation was unaffected when hemodilution was performed with either saline (r = 0.999) or lactated Ringer's (r = 1.000). The H/H results obtained with the iSTAT-1 instrument gave slightly less correlation with those of the GenS analyzer (r = 0.978 - 0.980) over this protein range. However, the iSTAT-1 results were generally lower than the GenS results, with discrepancies up to 2 g/dL for hemoglobin values and up to 4% for hematocrits at the lowest protein concentration. Therefore, it is recommended that H/H testing in patients with suspected hypoproteinemia or substantial hemodilution should be tested with a non-conductivity-based method.     

The relevance of a simultaneous measurement of WBC and CRP as a POCT(point of care testing) of the inflammatory markers running title: the POCT(point of care testing) of WBC and CRP

The POCT(point of care testing) is not enough popular as the term in Japan, but many tests are actually performed as POCT. We have developed POCT analyzer to measure WBC and CRP simultaneously from whole blood, and one of its key technologies is Hct correction of CRP concentration from whole blood to plasma. After its launch, the users are discussing some findings, one of what is a transition phase of WBC and CRP found by frequent measurement of both. In case of acute infectious inflammation, the CRP peak follows the WBC peak after 1 or 2 days. Needless to say, this finding makes haste to foresee a remission by both doctor and patient family. Thus, the POCT has a possibility to give additional and/or another advanced medical relevance even if it is alternative.     

Field study of lyophilised plasmas for local prothrombin time calibration in The Netherlands.

AIM: To assess the effect of a lyophilised calibrant plasma procedure on the international normalised ratio (INR) and its interlaboratory variation. METHODS: INR equivalent values were assigned to five lyophilised plasmas (one from normal donors and four from coumarin treated patients) by a reference laboratory using three calibrated thromboplastin reagents. The calibrant plasmas and five artificial control blood specimens were mailed to 44 Dutch laboratories for prothrombin time (PT) determination. The assigned INR values were used to calculate calibration lines for each participant laboratory. The calibration lines were then used to translate the PT of the control specimens to INR. RESULTS: For all lyophilised plasmas and control blood samples, there were significant differences between INR values determined with the three thromboplastin reagents. These differences could not be explained by inaccuracy of the international sensitivity index or mean normal PT of the reagents and must, therefore, have been induced by the preparation procedures for the lyophilised plasmas and control blood samples. The interlaboratory variation of the INR obtained with the calibrant plasma procedure had a coefficient of variation (CV) ranging between 2.1% and 7.3% and tended to be lower than the interlaboratory variation found with the usual methods (3.0-12.2% CV). There was a good agreement between the mean INRs obtained with the calibrant procedure and those obtained using the normal methods. CONCLUSIONS: The present study highlights the limitations of some lyophilised plasmas and control blood samples. It is not possible to assign a single INR value to each of these lyophilised plasmas and control specimens that is valid for all thromboplastin reagents. Nevertheless, by using reagent specific INR equivalent values for the calibrant plasma procedure, the interlaboratory variation could be reduced.     

Recombinant and tissue extract thromboplastins for determination of international normalised ratio in over-anticoagulated patients

The international normalised ratio (INR)/international sensitivity index (ISI) system is established for calibration of thromboplastins for laboratory monitoring of oral anticoagulant therapy. The calibration procedure employs patients stabilised on oral anticoagulants, and is therefore validated for patients within the therapeutic range. For practical reasons, the system is used for patients at all levels of therapy, including over-anticoagulated patients with particularly low levels of factors II, VII and X. We studied patients within and above the therapeutic range, using a thromboplastin containing recombinant human tissue factor (Innovin) and two tissue extract thromboplastins. In samples with INRs from 2.0 to 4.0, there was good agreement between results obtained with the three systems (mean INRs within 4% of each other). In patients with INRs > 4.0, results with a human placental extract reagent (Thromborel S) were similar to those obtained with a rabbit brain thromboplastin (IL PT Fib Hs Plus); mean INRs were 6.30 and 6.32 respectively (not significant). Results with Innovin (mean INR: 7.67) were significantly (P < 0.001) greater (on average by 22%) than those obtained with the other two materials. The discrepancy between results with different reagents negatively correlated with factor VII levels. Thus, the lower the factor VII level, the greater was the discrepancy between INRs. Unexpectedly, there was a positive correlation between factor V level and the difference between INRs with different reagents. Thus, the higher the factor V level, the greater was the discrepancy between INRs. The effect of these differences at higher INRs on patient management is unknown, but the recently revised UK guidelines recommend that management of these patients should be influenced by clinical factors, reducing the relative importance of discrepancies between results obtained with different systems.