International Reference Preparation For Thromboplastin Research Articles

Erratum

In the article by van den Besselaar et al. “A comparison of INRs determined with a whole blood prothrombin time device and two international reference preparations for thromboplastin” published in Thrombosis and Haemostasis in 2000 (Thromb Haemost 2000; 84: 410–412), the surname of the first author was incorrectly spelled as “van den Besselar”. The correct spelling is “van den Besselaar”.

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.

A Comparison of INRs Determined with a Whole Blood Prothrombin Time Device and two International Reference Preparations for Thromboplastin

Oral anticoagulant therapy is usually monitored with the prothrombin time (PT) on citrate plasma samples. In recent years instruments have been developed for measurement of the PT in non-citrated whole blood. In the present study, the manufacturer's calibration of one type of device (CoaguChek) in terms of the international normalized ratio (INR) was evaluated by one laboratory. Three subsequent lots of test strips for the CoaguChek were investigated using blood samples from 56 coumarin-treated patients. Citrated plasma samples from the same patients were analysed with two international reference preparations for thromboplastin (IRP), i.e., rTF/95 (recombinant human) and RBT/90 (rabbit brain). There were statistically significant INR differences between CoaguChek and the international reference preparations (p <0.001), but the mean relative deviation of the INR was not greater than 0.104. Clinically relevant criteria were used to assess the agreement between the CoaguChek and the IRP results. Standard agreement ranged from 82% to 95%. It is concluded that these test strips achieved a clinically acceptable level of accuracy. Further studies of patient management with these strips are justified.

Calibration of Local Systems with Lyophilized Calibrant Plasmas Improves the Interlaboratory Variability of the INR in the Italian External Quality Assessment Scheme

Calibration with lyophilized calibrant plasmas certified in terms of PT with International Reference Preparations for thromboplastin has been proposed to minimize the effect of coagulometers on the INR. Aim of this study was to test the ability of local calibration with lyophilized calibrant plasmas, combined with a modified statistical approach, to improve the interlaboratory variability of the INR measured on two test plasmas (one coumarin and one artificially-depleted) by participants in the External Quality Assessment Scheme (EQAS). Sets of lyophilized calibrant and test plasmas were sent to the participants in the EQAS, who were asked to determine PT with their own reagent/ instrument combination (local system). Results were returned as PT together with information on the type of local system, the stated International Sensitivity Index (ISI) and the geometric mean of PTs determined by testing with the local system fresh plasmas from 20 healthy subjects. Ninety-two participants using 9 and 11 brands of reagents and instruments returned results. The CV of the INR determined with the stated ISI for the coumarin (Mean INR = 4.39) and artificially-depleted (Mean INR = 4.23) test plasmas were 11.2% and 10.3% and were reduced on the average by 34% and 54%, respectively, when the INR was calculated with the local ISI. In conclusions, results from this field study involving laboratories and testing systems representative of the real situation in oral anticoagulant monitoring in our country, indicate that local calibration by artificially-depleted plasmas, combined with the proposed statistical approach, is suitable to improve the interlaboratory agreement on the INR.

Local Calibration of International Normalised Ratio Improves between Laboratory Agreement: Results from the UK National External Quality Assessment Scheme

In the present study we have performed local calibration of International Normalised Ratio (INR) measurement systems in a large series of laboratories. We assigned INRs to five lyophilised plasma calibrants, one prepared from normal plasma and four using plasma from warfarinised patients, using different International Reference Preparations for Thromboplastin. These five calibrants, and two lyophilised test plasmas were analysed by 349 centres using 60 different thromboplastin instrument combinations. Plasma calibrants were assigned INRs using the WHO reference thromboplastin RBT-90 or the European reference thromboplastin CRM 149R. Each participating centre determined PTs of the calibrants with their local system. These PTs were then used to construct a local calibration graph relating PT to INR. The PTs of test plasmas were converted directly into INR using the local calibration model and into INR using the conventional method. The overall medians of conventionally derived INRs of two test plasmas analysed in 349 centres were 2.50 and 3.10, compared to 2.47 and 3.04 after local calibration where RBT-90 was employed to assign INRs to calibrants. Use of CRM 149R to assign INRs to calibrants led to a significant (p<0.0001) increase in INR to 2.7 and 3.36 respectively. When results were grouped according to the thromboplastin employed, agreement between results with different reagents was improved by local calibration. There was a significant reduction (p<0.01) in the spread of results in different centres as indicated by a reduction in coefficient of variation.

Successful Calibration of a Reagent from Human Placenta against rTF/95, the WHO International Reference Preparation for Thromboplastin, Human Recombinant, Plain

Successful Calibration of a Reagent from Human Placenta against rTF/95, the WHO International Reference Preparation for Thromboplastin, Humen Recombinant, Plain -

Multi-Center Calibration of the Third BCR Reference Material for Thromboplastin, Rabbit, Plain, Coded CRM 149S

In a collaborative study by eleven laboratories performed within the framework of the European Community Bureau of Reference (BCR), a third reference material for thromboplastin, rabbit, plain, has been calibrated against RBT/90, the current WHO international reference preparation for thromboplastin, rabbit, plain. This third reference material (coded CRM 149S) has a mean International Sensitivity Index (ISI) of 1.257 with a standard error of the mean of 0.013. The previous BCR reference material for thromboplastin, rabbit, plain (coded CRM 149R) and the reference material for thromboplastin, bovine, combined (coded OBT/79) were also included in the trial for assessment of the long-term stability of the ISI values. The relationship between ISI values determined in the present study was nearly identical to that of the historical values. These results offer reassurance with regard to the long-term stability of these reference materials.

Multi-Center Study of Replacement of the International Reference Preparation for Thromboplastin, Rabbit, Plain

A multi-center study of the three W.H.O. international reference preparations (IRP) for thromboplastin has been performed. The main purpose of the study was the determination of the mean international sensitivity index (ISI) of a proposed replacement IRP for thromboplastin, rabbit, plain. Twenty laboratories in Europe, North America, and Australasia participated in the study. All clotting times were determined with the manual technique. The inter-laboratory variation of the ISI was considered as the relevant parameter for assessment of ISI precision. Low interlaboratory variation was observed in the calibration of rabbit thromboplastin against rabbit thromboplastin (2.9-3.0% coefficient of variation of calibration line slope). Relatively high interlaboratory variation was observed in the calibration of rabbit against human thromboplastin (3.7-7.2 CV). Intermediary interlaboratory variation was obtained in the comparison of rabbit or human plain thromboplastin with bovine combined thromboplastin (3.5-4.0% CV). These findings confirm the principle of like-to-like calibration in biological standardization. The mean ISI values based on the IRP for rabbit thromboplastin (RBT/79) are in excellent agreement with those obtained with the IRP for bovine thromboplastin (OBT/79). In contrast, the mean ISI values determined with the IRP for human thromboplastin (BCT/253) were 6% lower than those with RBT/79. This bias may be caused by a difference in design of the two historical calibration studies. The mean ISI of the proposed replacement IRP for thromboplastin, rabbit, plain obtained by calibration against RBT/79 was 1.035 (standard error: 0.027). It is recommended to round the mean ISI off to 1.0. Opportunity was also taken to compare a lyophilized normal pooled plasma with fresh individual normal plasmas.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of the choice of WHO International Reference Preparation for thromboplastin on International Normalised Ratios.

To compare the International Normalised Ratio (INR) obtained directly with the two types of WHO plain International Reference Preparation for thromboplastin in patients treated with coumarin. Prothrombin times were performed in parallel at four centres using WHO human plain IRP (BCT/253) and rabbit plain IRP (RBT/79). Sixty patients and 20 normal controls were tested at each centre. Differences in INR among the centres were assessed by one factor, analysis of variance. The bias for each centre was assessed by the t test. At all four centres higher INRs were consistently found with the rabbit plain reagent. Two of the centres showed significantly greater bias. There was a small but significant difference in INR results obtained directly with these two reference reagents at all four centres (mean 7.35%). This in part may result from the different responsiveness of the two IRP to the coumarin defect or to imprecision of the original ISI calibrations of the two plain WHO IRP. The findings support the adoption of a single master IRP, in accord with WHO recommendations, which would resolve the present anomalous situation.

Methodische Aspekte zur photometrischen Bestimmung der Prothrombinzeit mittels chromogener Substrate

A method for photometric determination of prothrombin time (PT) with a chromogenic peptide as substrate is described. The reagent contains human placental thromboplastin, a chromogenic substrate, calcium, a heparin antagonist and buffer. The new prothrombin time method has been calibrated against international reference preparations for thromboplastin. The reagent is sensitive to deficiency of all coagulation factors of the extrinsic pathway. However, it is not sensitive for heparin up to 1 IU/ml. The precision of this fast and simple method is comparable to that of mechanised assays for clinical chemistry.

Standardization of the prothrombin time in oral anticoagulant control.

The establishment of International Reference Preparations for thromboplastin together with a recommended methodology for use has permitted the definition of a universal scale for intensity of oral anticoagulation. This scale is called International Normalized Ratio (INR). INRs can be calculated by using calibrated thromboplastin/instrument systems. A calibration model has been developed together with statistical methods to test this model. The imprecision of calculated INRs has been estimated using information from international collaborative studies and proficiency testing programs. Normalized prothrombin times obtained with calibrated thromboplastin/instrument systems have been shown to provide safe patient care in clinical practice. The development of control plasmas with assigned INR equivalents will complete the standardization system. The INR scale will facilitate the process of consensus-making with respect to optimal therapeutic ranges for anticoagulant intensity. Finally, it will improve the continuity of anticoagulant control of travelling patients.

The Calibration of the Second Primary International Reference Preparation for Thromboplastin (Thromboplastin, Human, Plain, Coded BCT/253)

An international collaborative exercise has been undertaken to calibrate a replacement for the first WHO primary international reference preparation (IRP) for thromboplastin. The replacement preparation is a lyophilised batch of British Comparative Thromboplastin (BCT/253, human plain) for use in the Quick prothrombin time test. Seventeen centres participated. The experimental design, calibration model and statistical analysis were based on the recommended WHO procedure. As a result of this calibration exercise an International Sensitivity Index (ISI) of 1.1 has been assigned to the preparation by WHO and it has been officially recognised as the second primary IRP for thromboplastin. The calibration of BCT/253 is an essential link in a new hierarchical structure for the standardisation of the prothrombin time. The aim is to provide a uniform international system of reporting prothrombin time results using International Normalised Ratios (INR) derived from the ISI of individual thromboplastins.