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Prothrombin Time and INR

The prothrombin time (PT or protime) is the actual time, measured in seconds, for an anticoagulated sample of patient plasma to clot after the addition of calcium and an activator of the extrinsic pathway, which is called thromboplastin. The PT assesses the coagulation factors of the extrinsic pathway (factors VII) and the common pathway (activated factor X, activated factor V, prothrombin and fibrinogen). PT reagents are more sensitive to Factor VII deficiency than to deficiencies of factors V, X, prothrombin and fibrinogen. 

Heparin neutralizers, such as Polybrene, are added to PT reagents and can neutralize up to 2 U/mL of heparin. However, the presence of excessive heparin (sample collected shortly after a heparin bolus, collected above the site of a heparin infusion or from a heparin coated catheter) may overwhelm the neutralizer and prolong the PT because of its inhibitory effect on thrombin. 

Physicians may request a PT any of the following reasons: 

  • Investigate a history of a bleeding tendency prior to invasive surgical procedures 
  • Prior to intravascular cardiac surgery or other procedures that interfere with coagulation 
  • Determine the etiology of abnormal bleeding 
  • Assess liver function. 
  • Monitor warfarin therapy

The prothrombin time may be prolonged by deficiency of a single or multiple coagulation factors (Factor VII, Factor X, Factor V, Factor II or fibrinogen).  The prothrombin time may also be prolonged due to the presence of a coagulation inhibitor such as a specific factor inhibitor or the lupus anticoagulant.  

Warfarin acts in the liver by inhibiting the synthesis of vitamin K dependent clotting factors, which include factors II, VII, IX and X, and other proteins essential for the clotting process. Warfarin starts working in the liver within 24 hours, but the maximum effect of a single dose may not be seen for 2-4 days. If administered on a daily basis, it usually takes 5-10 days for the warfarin level in the patient’s bloodstream to reach a steady state. Due to significant patient variability in response to warfarin therapy, the PT/INR must be closely monitored until a steady state has been reached. Typically, monitoring is performed daily, until several days after the therapeutic range is reached. Warfarin dosage changes may also be required in response to the PT/INR results. However, until the patient reaches a steady state, PT/INR fluctuations are expected. It may take up to one month for a physician to reach an optimal therapeutic level of warfarin for an individual patient. 

One common cause of vitamin K deficiency is chronic administration of broad-spectrum antibiotics, which decreases the intestinal flora that synthesize vitamin K. Examples include cephalosporins, ciprofloxacin, erythromycin, ofloxacin, sulfonamides, tetracyclines neomycin, chloramphenicol and metronidazole. Antifungal medications such as fluconazole, miconazole, norfloxacin and itraconzole can also prolong the PT. 

Other medications can affect the action of warfarin in the liver. Analgesics including acetaminophen, aspirin and ibuprofen can prolong the PT. Other examples include anticonvulsants, omeprazole, cimitidine, simvastatin, propylthiouracil, urokinase, streptokinase, amiodarone, thiazides, tricylclic antidepressants, danazol, and allopurinol. 

Other drugs can antagonize the effect of warfarin. Examples include estrogen, corticosteroids, barbiturates, carbamazepine, spironolactone and some antibiotics such as dicloxacillin, nafcillin and rifampin. 

 Any time a physician changes or adds medication to a patient’s regimen, the PT/INR should be checked to ensure that the patient maintains the proper level of anticoagulation. Alcohol, changes in diet, infections, liver disease, and other illnesses also can alter the patient’s PT/INR. Periodic monitoring, up to once every month, is necessary as long as the patient remains on anticoagulation therapy. More frequent routine monitoring may be required in some patients. 

The American College of Chest Physicians and the National Heart Lung and Blood Institute revised their recommendations for intensity of warfarin therapy in 1995. An INR of 2.0 to 3.0 is recommended for all indications except mechanical prosthetic heart valves, for which an INR of 2.5 to 3.5 is recommended.     

Recommended Therapeutic Ranges for Oral Anticoagulant Therapy



Prevention of venous thrombosis

Treatment of venous thrombosis

Treatment of pulmonary embolism

Prevention of systemic embolism

                     Tissue heart valves

                     Acute myocardial infarction

                     Valvular heart disease

                     Atrial fibrillation

2.0 – 3.0

Mechanical prosthetic valves

2.5 – 3.5


While the PT is a common test for evaluating the coagulation of the patient, differences in thromboplastin reagents have caused problems when comparing results across laboratories. This lack of comparability is of special concern for patients who may use more than one laboratory for PT testing.  This has been due to varying sensitivities of different thromboplastin reagents used in the PT test.  The World Health Organization has recommended a scheme for standardization of oral anticoagulant monitoring, based on expressing PT results in terms of an International Normalized Ratio (INR).  

Various thromboplastin reagents have different sensitivities to the warfarin-induced reduction in coagulation factor synthesis. Since 1983 the International Normalized Ratio (INR) has been used to standardize PT results for patients on warfarin.  The INR is a calculation designed to minimize the variability of PT results due to differences in sensitivity of thromboplastin reagents. With the INR, results from different laboratories utilizing different methodologies can be compared. Even with the INR, however, significant interlaboratory variations in INR results have been reported. 

The international sensitivity index (ISI) reflects the sensitivity of the reagent as compared to an international standard. The manufacturer of the thromboplastin reagent determines the ISI by comparing each batch of reagent to a World Health Organization reference plasma and then assigning an ISI value to that lot of reagent. Reagent and instrument specific ISIs reduce the variability in INR results. The INR is calculated from the following formula: 

INR = (PT patient / PT normal )ISI 

• PT patient is the patient’s PT result expressed in seconds. 

• PT normal is the laboratory’s geometric mean value for normal patients expressed in seconds. 

Each laboratory must determine the normal range for the patients it serves in order to calculate an accurate INR. To find the normal range, a minimum of 20 non-anticoagulated healthy patients evenly distributed between males and females should be tested. Emergency department or pre-op patients should not be used for the normal patient pool, because their blood may contain acute phase reactants that are elevated in times of stress and inflammation and can shorten the normal range. Testing should be completed over a period of several days to include intra-lab variabilities.

The INR result is the patient’s PT result in seconds divided by the geometric mean of PT result of the laboratory’s normal patients, as calculated by each laboratory. The geometric mean is a type of average that is different from the simple arithmetic average. It is calculated by multiplying all the PT results together (in this case, the 20 normal PT results) raised to the reciprocal of the number of results (in this case, 1/20). The geometric mean is used to avoid bias that may be caused by the inclusion of extremely high or low values. A calculator or software program is necessary to calculate geometric mean. 

Because of the exponential nature of the equation for calculating the INR result, a lower ISI yields a more accurate result. The College of American Pathologists (CAP) Conference on Laboratory Monitoring of Anticoagulant Therapy held in 1998 recommended that an ISI between 0.9 and 1.7 be used, with the desired range being at the lower range of this scale. The following table illustrates the effect that the ISI can exert on the INR result. 

Impact of ISI on INR Results

Patient PT result 


Calculated INR result 











INR calculations are intended to yield identical INR results when a single specimen is tested by two laboratories: one using a more sensitive thromboplastin (yielding a higher PT result) and the other using a less sensitive thromboplastin (yielding a lower PT result). 

It was previously believed that INR precision would be improved by using more sensitive thromboplastins (with lower ISI values). This has been shown to be incorrect. One intrinsic source of error in INR reporting is the fact that any imprecision in the PT measurement is amplified exponentially when the INR is calculated. This imprecision is greater for higher PT and INR values. Other problems contributing to lack of INR reproducibility include differences in sample collection or handling, and inherent biological variation in individuals. All of these limitations in the INR system are illustrated by a study in which parallel INR determinations were performed at three different hospitals within a single community, using split plasma samples from patients on long-term warfarin therapy. When the patient INR’s were classified as therapeutic, sub- or supratherapeutic, interlaboratory discrepancies were observed in 30% of paired samples, with the majority of discrepancies occurring with INR values greater than 3.0. 

Laboratories must do extensive comparison studies before switching to a new lot of PT reagent or changing methodology to confirm accuracy of the assigned ISI value. The CAP Coagulation Resource Committee has published a checklist to assist laboratories with prevention of problems with the INR calculation.

• Has an appropriate reference range for your current reagent/instrument been identified? 

• Is the geometric mean of this reference range being used in the calculation? 

• Is the correct ISI being used in the calculation? The ISI varies not only with the lot of reagent but also with the instrument being used. 

• When performing the calculation, the values for the patient PT and the PT reference range mean include one decimal place (e.g. 12.0) and ISI includes two decimal places (e.g.1.05). In each case, three significant digits are used. The INR should then be rounded and reported to one decimal place (e.g. 2.5; two significant digits). 

• How is the calculation being performed? Manually? By the instrument? By the LIS? The calculation should only be performed by one of these methods. Having more than one method in place increases the maintenance efforts and increases the risk of miscalculation. 

• If a manual method of calculation is used, how do you minimize errors? 

• Is the calculation tested periodically to assure that the correct INR is being produced? In particular, this calculation must be tested immediately after changing any component of the formula (ISI or the geometric mean of the reference range). Actual patient reports should be periodically reviewed to assure the appropriately calculated result is being reported. 

• Are specimens collected into 3.2% citrate-not 3.8%? Published reports have documented that INR results differ between the two citrate concentrations. Some laboratories continue to use 3.8% citrate despite international efforts to standardize on 3.2%. 

One important practical issue related to the imprecision of the INR is the question of warfarin dose adjustment. Random variation of INR values may occur in a patient on stable oral anticoagulant dosage, as a result of both biological and analytic variation. This total random variation has been estimated at ~10%. This data has been used to evaluate whether a change in the INR represents clinically insignificant random variation, or a clinically relevant change requiring warfarin dose adjustment. It has been calculated that in a patient on fixed dose and steady state warfarin, a change in the INR is significant only if it is a change (increase or decrease) of greater than 0.28 times the previous INR value. This is detailed in the accompanying table.

Previous INR

Significant change in INR would be greater than

Range of acceptable INR variation



1.4 - 2.6



1.8 - 3.2



2.2 - 3.8



2.5 - 4.5


In summary, the INR reporting system is a major improvement over PT values for standardization of oral anticoagulant monitoring, however clinicians should bear in mind that the system is not problem-free, and that apparent discrepancies may arise between different laboratories, and over time within a single patient.

The College of American Pathologists recommends using a sensitive thromboplastin reagent for measuring prothrombin times (PT’s). The reason is that more sensitive reagents are associated with improved precision in PT testing, as well as a wider therapeutic PT range, resulting in more reliable monitoring of oral anticoagulant therapy, and facilitation of dose adjustment.

Preanalytic Variables 

There are many variables that can affect the PT/INR result outcome. In fact, preanalytical variables account for up to 64% of all errors in PT/INR testing. One of the most important factors is the anticoagulant used in the blood collection tubes. The World Health Organization and NCCLS guidelines recommend the use of 3.2% trisodium citrate.  This anticoagulant chelates calcium, preventing activation of the coagulation system. The evacuated tube must be completely filled (at least 90% full) to maintain the proper 5:1 plasma to anticoagulant ratio. If a patient’s hematocrit is >55%, volume of plasma collected is proportionally decreased and the volume of anticoagulant increased, resulting in artifactual prolongation of the PT. A similar problem occurs when the blood collection tube is underfilled.  

 It is best to avoid traumatic venipunctures to minimize the release of tissue factor, which can initiate coagulation.  Although not critical for coagulation specimens, it is always good practice to release the tourniquet as soon as possible. The blood sample must be centrifuged for sufficient time (10 minutes) to create platelet-poor plasma as the presence of platelets in the specimen can shorten clotting times. Specimens for PT testing may be stored at room temperature and will yield valid results for specimens stored up to 24 hours, provided that the collection tube remains unopened. If testing cannot be performed within this time period, the platelet-poor plasma should be frozen. 

Reference range is 11.9–14.3 seconds. Critical values are an INR >2.0 in patients who are not anticoagulated and an INR of 4.0 if the patient is anticoagulated. 

Specimen requirement is one light blue top (sodium citrate) tube of blood drawn after a 5 mL red top discard tube.

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