Problems In Laboratory Monitoring Of Heparin Dosage

Heparin is a sulphated polysaccharide widely used as an anticoagulant for the treatment and prevention of thromboembolic disease (Hirsh, 1991; Hyers et al, 1992; Hirsh Hirsh Abildgaard, 1968). After heparin binding, AT undergoes a conformational change and is converted into a very rapid inhibitor with the active centre arginine residue in AT much more reactive against the active centre serine of several coagulation enzymes (Lindahl et al, 1979; Rosenberg Kijowski et al, 1994). A number of different blood sample collection systems have been used for collection of blood samples for heparin monitoring that vary in respect of tube composition (plastic or glass), and siliconization and anticoagulant. There is evidence that use of 0·109 mol/l trisodium citrate (3·2%) or 0·129 mol/l trisodium citrate (3·8%) can influence the activated partial thromboplastin time (APTT) determinations in samples from patients receiving intravenous UFH, results being on average 10% greater in samples collected into 0·129 mol/l citrate (Adcock et al, 1997). The lower strength is increasingly used as it is recommended for samples collected for monitoring oral anticoagulant therapy (WHO, 1999). It is now well known that, after blood has been collected, any heparin present may be partially neutralized during any delay before analysis. This is caused by release of platelet factor 4 (PF4) from platelets within the sample (Rucinski et al, 1979). The PF4 binds to heparin and neutralizes it. In order to combat this problem, samples from heparinized patients can be collected into an anticoagulant cocktail that inhibits platelet release, thereby avoiding PF4-mediated neutralization of heparin. It has been demonstrated that collection of blood into a citrate, theophylline, adenosine, dypyridamole mixture (CTAD) can greatly reduce the neutralization of heparin activity in blood (Contant et al, 1983; Van den Besselaar et al, 1987). In the study of Van den Besselaar et al (1987), APTTs of blood samples collected into CTAD were associated with less than 11% shortening depending on APTT reagent, compared with a 15–29% shortening of citrated APTTs between 1 h and 5 h after collection. Based on their results, the authors recommended the use of CTAD for heparin monitoring samples. The same study concluded that room temperature was the most appropriate storage temperature and also recommended minimum centrifugation at 940 g for 30 min or 2200 g for 10 min. These studies suggest that if the APTT is used for heparin monitoring, it is important to perform the measurement within 5–6 h if CTAD is used as the anticoagulant. If citrate is used, testing of samples more than 2 h after collection may lead to important underestimation of in vivo heparin effects. Freezing samples for later analysis should be avoided as this can lead to shorter APTTs unless great care is taken to remove the maximum possible number of platelets (Van den Besselaar et al, 1990). There is evidence that the volume of blood within a particular vial size can influence the APTT as determined for UFH control, even when the ratio of anticoagulant to blood (1:9) is maintained (Ray, 1991; Ray et al, 1993). The mean APTTs of 5 ml samples from heparinized patients was 120 s compared with a mean of 79 s for 2 ml samples (Ray, 1991) when both were collected into containers with a capacity of 6·7 ml. The author speculated that the air bubble of 4·7 ml within the sample containing 2 ml of blood caused greater mixing, release of PF4 and neutralization of heparin than the 1·7 ml air bubble in the samples containing 5 ml of blood. In a subsequent study, increased release of PF4 produced similar effects (i.e. significantly shorter APTTs in the container with a smaller volume of blood) if citrate was used, but not if CTAD was used, as the anticoagulant (Ray et al, 1993). For heparin monitoring it is therefore inappropriate to use sample containers that contain a large air volume unless CTAD is used as the anticoagulant. The timing of blood sample collection for heparin monitoring should depend on the administration of the drug. If heparin is given by intravenous infusion, the sample should be collected after equilibrium has been reached (Samama, 1995a), normally 4–6 h after initiation of the infusion. A similar lag phase should pass prior to sampling following a dose adjustment. When heparin is given by subcutaneous injection, the maximal response of the APTT occurs after 4–6 h (Griffith Turpie et al, 1989). There are conflicting reports in relation to possible diurnal changes in the anticoagulant effect of infused heparin. In studies of heparinized patients (Decousus et al, 1985) or healthy volunteers (Krulder et al, 1992) there was considerable diurnal variation, with significantly greater APTT prolongation in the night (3·30–8·30am) than in the day (Krulder et al, 1992) and up to 60% difference in individual results between night and day (Decousus et al, 1985). Such a diurnal effect was not detected in two further studies (Toulon et al, 1987; Fagrell et al, 1989). On occasion, it may be necessary to eliminate heparin in a plasma sample. Several methods for removal of heparin from plasma have been described that involve absorption of heparin by cellulose resin (Cumming et al, 1986; Wenz et al, et al, particularly factor As factor is an levels probably vary in a of patients with thromboembolic disease and this may have to the difference in of described in some of the There are a number of that to between in respect of heparin including the use of different and In there is of the of APTT used in the et al, and in the (Kitchen et al, The of APTT a range from (Kitchen et al, was from a from material and from in containing (Kitchen et al, It is therefore not that as influence heparin et al, 1986; Van den Besselaar et al, 1993). In study (Kitchen et al, there was a between a lower of present as or and increasing heparin There is also evidence that different with the same are associated with in heparin Turpie et al, et al, 1994). The evidence for an increased risk of bleeding when a heparin of is is less well (Hull et al, although there are reports of increased bleeding at levels of or greater by et al, to the of between in respect of heparin is for a to a range for the APTT in use that to a heparin of by or by (Hirsh, 1991). As the between heparin and APTT is different to heparin is to plasma in compared with this must be samples from patients receiving heparin. This has been used in at least studies different APTT et al, et al, et al, have compared the results of five and clotting in samples from patients receiving or and between the results of different techniques (Kitchen et al, of the clotting were significantly lower than results by some other clotting and by in therapy and in a which was by the of with different methods of (Kitchen et al, It has been suggested that the should be the of et al, of the influence of activity on some clotting although not clotting techniques are in this et al, et al, There is evidence that the of results in different samples from patients receiving different LMWH preparations be reduced if LMWH was used in the of in of heparin et al, and there is evidence that the for heparin is as a for LMWH et al, 1985). The of for of for monitoring LMWH therapy should therefore be a LMWH that has been against the for LMWH et al, 1988). A number of have now been that heparin monitoring at the Some of are for during or et al, have been for monitoring patients with heparin for thromboembolic disease et al, et al, et al, 1996). have the of results from the requirement for centrifugation of samples. blood collected, for by required blood. UK for testing in including monitoring of heparin have been et al, and testing of samples to the range of heparin levels in the being compared to the APTT in use and to activity. These that the of the should for of the with of results, and that should be by an appropriate of the may also for The and of or testing in coagulation have been of the most important of heparin therapy is in platelet in as many as 10% of patients receiving heparin and to during therapy The reduction in platelet to as may be associated with and between the and day after initiation of therapy in patients should be if to of the initial platelet after several of heparin therapy As UFH therapy is if occurs and can be successfully used in some et al, patients receiving heparin platelet to be in to monitoring, particularly therapy is It has been suggested that patients receiving heparin for the time should have platelet at and on after day monitoring is required from the in patients with to heparin et al, The anticoagulant of for monitoring UFH therapy is CTAD (Van den Besselaar et al, 1987). Samples should be collected 4–6 h after initiation on infusion, injection or dose et al, When employing APTT for monitoring UFH, the of the monitoring must be taken into when a range et al, & and should to by or by (Hirsh, 1991). The heparin can be used for monitoring UFH, particularly when APTT is inappropriate et al, 1994). patients receiving LMWH as or patients for venous thromboembolism by a do not monitoring et al, LMWH may monitoring in as therapy including at risk of bleeding or and patients with or low et al, A range of has been recommended for patients receiving twice LMWH therapy et al, for LMWH should be a material against the for of may vary to the (Kitchen et al,