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Background: Prothrombin time (PT) has long been the most popular test for monitoring oral anticoagulation therapy. The International Normalized Ratio (INR) was introduced to overcome the problem of marked variation in PT results among laboratories and the various recommendations for patient care. According to this principle, all reagents should be calibrated to give identical results and the same patient care globally. This is necessary for monitoring of single patients and for application of the results of anticoagulation trials and guidelines to clinical practice., Methods: We took blood samples from 150 patients for whom oral anticoagulation had been prescribed. Plasmas were separated and PTs determined by use of seven commercial reagents and four calibrator sets. The differences in results were assessed by plotting, for each possible pair of methods, the differences in INR values for each sample against the mean INR value (Bland-Altman difference plots)., Results: Mean results differed significantly (P <0.001) for 17 of 21 possible paired comparisons of methods. Only two pairs of methods produced very similar results when assessed for problems of substantial differences in INR values; a significant, systematic increase in the difference with INR; and a significant systematic increase in the variation in difference with increasing INR values., Conclusions: The agreement among several (and perhaps most) commercial INR methods is poor. The failure of current calibration strategies may severely compromise both the monitoring of individual patients and the application of oral anticoagulation guidelines and trial results to clinical practice.

Background: For monitoring of treatment with oral anticoagulants, the clotting time obtained in the prothrombin time (PT) test is transformed to the International Normalized Ratio (INR) with use of a system-specific International Sensitivity Index (ISI). The calibrant plasma procedure (CPP) is an alternative approach to INR calculation based on the use of a set of lyophilized plasmas with assigned INRs., Methods: With the CPP, a linear relationship is established between log(PT) and log(INR), using orthogonal regression. CPP was validated for Simplastin HTF, a new human tissue factor reagent derived from cultured human cells. CPP precision was assessed as the CV of the slope of the regression line. The accuracy of the CPP was determined by comparing the INR obtained with the CPP with that obtained with the established ISI-based reference method. INRs of the calibrants were assigned by different routes: by manufacturer (consensus labeling) or by use of Simplastin HTF or International Reference Preparations (IRPs; rTF/95 or RBT/90)., Results: The mean CV of the CPP regression slope ranged from 1.0% (Simplastin HTF reagent-specific INR) to 2.4% (INR assigned with rTF/95). INRs calculated with the CPP were similar to those obtained with the reference method, but when the routes for assigning INRs to the calibrant plasmas were compared, the mean difference in INR between CPP and the reference method was smaller with Simplastin HTF reagent-specific values. In several (but not all) cases, this difference was significant (P <0.05, t-test)., Conclusion: CPP can be used for local INR determination, but better precision and accuracy are obtained with reagent-specific INRs compared with INR assignment by consensus labeling or IRP.

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.

Background: The International Sensitivity Index (ISI) calibration of whole-blood prothrombin time (PT) monitors for point-of-care testing (POCT) described by Tripodi et al. (Thromb Haemost 1993;70:921-4) has been shown to be dependable but is too complex and demanding. The use of plasma would simplify calibration of whole-blood POCT PT monitors, but important differences may exist between the ISI for whole blood and plasma calibrations., Methods: In a 10-center calibration study of two POCT whole-blood monitoring systems (CoaguChek Mini and TAS PT-NC), we characterized the relationship between the log PT for whole blood and fresh plasma with use of single lots of test strips/cards. This relationship (linear) was used to correct the difference between the whole-blood and plasma ISI. The reliability of the correction with different lots of test strips/cards was assessed at three centers. The linear relationship was used to correct the difference in the whole-blood and plasma ISI with four other lots of TAS PT-NC cards and with two additional lots of CoaguChek Mini test strips., Results: The correction decreased the ISI difference from 13.3% to 0.9% for the TAS PT-NC and from 5.7% to 0.6% for the CoaguChek Mini. In assessments at three centers, which included different lots of test strips/cards, the mean ISI difference was markedly decreased with the TAS PT-NC but not with the CoaguChek Mini, for which the mean ISI difference increased slightly., Conclusions: The proposed correction resolves the discrepancy between whole-blood and fresh plasma ISI calibrations with TAS PT-NC test cards. The CoaguChek Mini systems could be calibrated without this correction.

The international sensitivity index (ISI) calibration of point-of-care-test (POCT) prothrombin time (PT) whole blood monitors is complex, requiring manual PT testing of 60 patients' and 20 healthy subjects' plasma samples. The possibility of reducing these numbers was studied by a Monte Carlo Bootstrap study for 2 POCT PT systems. For reduced sample numbers, this consisted of 50,000 calibrations using whole blood and plasma samples tested on the monitors with manual PT testing of plasma samples from the same blood donations. There was little effect on mean ISI by reduction of sample numbers to a total of 7, but there was progressively less certainty regarding the reliability of the calibration. Precision of the calibrations and international normalized ratio deviation were not affected markedly by reducing numbers to half As ISI calibration with the 2 POCT systems was less precise than conventional manual testing, for maximum confidence, reduction of numbers is not advised.

Background: To simplify International Sensitivity Index (ISI) calibration, the possibility of substituting fresh plasma for fresh whole-blood samples with point-of-care testing (POCT) whole-blood monitors was investigated in a three-center study of three different POCT systems., Methods: A modified full WHO calibration procedure based on 20 healthy controls and 60 coumarin-treated patients was performed on three monitoring systems with whole-blood and plasma samples against plasma tested using the European Concerted Action on Anticoagulation (ECAA) rabbit reference plain thromboplastin and the manual prothrombin time (PT) method., Results: With one of the three systems, the mean ISI was 1.51 for whole blood and 1.49 for plasma; with the second system, the mean ISI was 1.08 for both whole blood and plasma. With the third system, however, the difference between the mean ISI for whole blood and that for plasma was greater (1.15 and 1.01, respectively). Overall, the precision of the calibrations was less than with traditional manual plasma PT testing., Conclusions: Provided that an appropriate calcium chloride concentration is used, the plasma PT results can be used for accurate ISI calibration of two of these three whole-blood POCT systems. Precision criteria need to be modified for POCT monitors.

The international normalized ratio (INR) was established as a means of standardizing the prothrombin time regardless of the thromboplastin used in the individual laboratories. The INR is the prothrombin time ratio of the sample raised to the power of the International Sensitivity Index (ISI). Traditionally, the ISI is determined by using a manual clotting technique by comparing the test thromboplastin with a World Health Organization international reference thromboplastin with results from 60 patient samples standardized on warfarin, and 20 samples from normal volunteers. Most laboratories no longer perform prothrombin time testing by a manual technique but instead automate the procedure by using a variety of coagulation instruments. Thromboplastin ISI values have the potential to be modified considerably by instrumentation, and as a consequence, they, may result in INRs that are unreliable when compared to those obtained by the traditional manual method. It has been proposed that the use of calibrant plasmas tested with the laboratory's thromboplastin and instrument may overcome this problem and a local ISI value could be established. Our study objective was to determine whether a local ISI calibration with the plasma calibrants could reduce the variation of the INR over a wide range of thromboplastin/ instrument combinations. A total of 58 laboratories from across Canada, including community hospitals, private laboratories, and large tertiary care units, participated. The findings indicate that the use of calibrant plasmas to calculate the local ISI does improve the accuracy of INR reporting in the majority of thromboplastin/instrument combinations.

A written survey of all licensed medical laboratories performing coagulation testing in Canada was undertaken in 1996 as a follow-up to a study performed in 1992. The an overall response rate was 77.4% of the laboratories surveyed. During the 4-year period between surveys, the percentage of responding laboratories using some format for international normalized ratio (INR) reporting increased from 57% to 98%. The mean International Sensitivity Index (ISI) of the various thromboplastins used had decreased from 2.07 to 1.63. However, only 35% of responding laboratories used a thromboplastin with an ISI of 1.2 or less as recommended by the American College of Chest Physicians. In 92% of cases, the ISI was said to be specified for the type of instrument, ie, optical density vs mechanical. However, in only 52% of cases was the ISI known to be specific for the particular reagent-analyzer combination used in the individual laboratory. Nine percent of the laboratories were still using an inappropriate denominator for the INR calculation. While 87% of laboratories quoted a therapeutic INR range of 2.0 to 3.0 for venous thromboembolism, there was much greater variation in the range reported for patients undergoing heart valve replacement. The situation in Canada has improved in using the INR system for prothrombin time reporting. However, additional education is required, as is greater use of thromboplastins with a lower ISI.

The comparative value of lyophilized artificially depleted plasmas and plasmas from warfarin-treated patients in local (thromboplastin/coagulometer) system International Sensitivity Indices (ISI) determination has been studied at 39 centers in an international collaborative study. At each center, both types of plasmas were tested with two common thromboplastins (low ISI RecombiPlasTin and high ISI OBT), and in-house reagents using three brands of coagulometers. In 95 local systems, they was agreement. With both thromboplastins, coagulometers lowered the ISI from the manual or stated ISI with all systems. Differences between the two types of lyophilized plasmas with the two common thromboplastins were not of clinical importance. Artificially depleted plasmas are easier to obtain in sufficient volumes than plasma from warfarin-treated patients, and more easily provide the wide range of INR values required for reliable local ISI calibration. The agreement between the two types of lyophilized calibrant is reassuring.

A study of lyophilized plasma calibrants in correcting for coagulometer effects on International Normalized Ratios (INR) has been conducted in an international survey. Prothrombin times were performed with the calibrants on 2 "common" thromboplastins and in-house thromboplastins in 3 brands of coagulometer at 37 centers. The International Sensitivity Indices (ISI) of 95 local systems with the calibrants were derived. The "true" INR of 10 test plasmas from warfarin-treated patients were established centrally using thromboplastin International Reference Preparations (IRP). International Normalized Ratios of these plasmas for each center were calculated using both the manufacturers' stated and the local ISI. The mean deviation of the 95 systems from the "true" INR of the warfarin plasmas was +14.4% with the manufacturers' ISI, but reduced to +1.04% with the local ISI. Local ISI determination with the calibrants avoids many of the difficulties of conventional thromboplastin calibrations. Plasmas from patients on warfarin and parallel manual PT with thromboplastin IRP are not required.

Prothrombin times accumulated from ten different proficiency testing surveys were analyzed in terms of a linear additive model described by Evatt et al (Clin Lab Haemat 1981; 3:331-342). Different types of lyophilized plasma samples were used, i.e., plasmas artificially depleted of coagulation factors by adsorption to aluminum hydroxide, and pooled plasmas of patients receiving coumarin drugs. For each plasma sample, both instruments and thromboplastins had a highly significant effect on the prothrombin time. For most instruments and thromboplastins, a good correlation was found between instrument effect or thromboplastin effect and the mean prothrombin time if various artificially depleted plasma samples from a single manufacturer were used. Artificially depleted plasmas from a second manufacturer gave different relationships between estimated effects and mean prothrombin time. Relationships based on lyophilized pooled patient plasmas were different from those of artificially depleted plasmas from either manufacturer. The potential use of the additive linear model for standardization of the prothrombin time in monitoring oral anticoagulant therapy is discussed. Additional studies are required to establish the suitability of this model to define a universal scale for prothrombin times of fresh plasma samples of anticoagulated patients. If suitable, the model can be linked to the International Normalized Ratio proposed by the World Health Organization.

Data from the 1976 and 1977 CAP surveys were analyzed for response of the activated partial thromboplastin time (APTT) to heparin. Different sources and concentrations of heparin were used. The results indicate that the precision of the APTT is more dependent on instrumentation than on partial thromboplastin. This was true for all four of the heparinized specimens evaluated. A single exception was found with the "old" Dade reagent activated cephaloplastin. The mean difference in the activated partial thromboplastin times obtained with differing concentrations of heparin was entirely dependent on the partial thromboplastin reagent used. No significant difference in the results was found when equal concentrations of bovine lung and porcine intestinal mucosal heparin were compared.

Data from the 1976 CAP Survey for the performance of Factor VII (antihemophilic factor) assays were analyzed. The results indicated an undesirable variation on both normal and abnormal specimens. Reagent-instrumentation systems and procedures were examined. The need for standardization of factor VIII assays is evident from the results of this study.