APTT assay

Anticoagulant Activity Assay. The anticoagulant activities of IV-acetylated heparin and all further derivatized heparins were determined based on activated partial thromboplastin time (APTT) assay methods (15). Bovine blood was collected in 3.8% sodium-citrated (9 parts blood to 1 part citrate solution) and centrifuged at 5000 x g for 15 min. The supernatant plasma was collected and pooled for subsequent APTT testing. Prior to testing plasma was kept refrigerated no longer than 6 h after... 

Activated partial thromboplastin time (aPTT) is a coagulation assay, which measures the time for plasma to clot upon activation by a particulate substance (e.g., kaolin) in the presence of negatively charged phospholipids. 

II Umbilical cord, joint, and mucosal tract bleeding Prolonged prothrombin (PT) and activated partial thromboplastin time (aPTT) Specific assay for prothrombin... 

V Mucosal tract bleeding Prolonged PT and aPTT Normal thrombin time Specific factor V assay... 

VII Mucosal tract, joint and normal aPTT Prolonged PT Specific factor VII assay... 

XIII Umbilical cord, intracranial, and joint bleeding recurrent miscarriages, impaired wound healing Normal PT, aPTT, thrombin time, bleeding time Specific factor XIII assay... 

The CTAD additive mixture has found application in the monitoring of heparin therapy by either the chromogenic substrate assay or the APTT and in the measurement of platelet markers such as P-selectin (CD62) by flow cytometry (108, 109). 

One molecule of lepirudin binds to one molecule of thrombin and thereby blocks the thrombogenic activity of thrombin. As a result, all thrombin-dependent coagulation assays are affected (eg, aPTT values increase in a dose-dependent fashion). Pharmacokinetics ... 

Lab test abnormalities In general, adjust the dosage (infusion rate) according to the aPTT ratio (patient aPTT at a given time over an aPTT reference value, usually the median of the laboratory normal range for aPTT see Administration and Dosage). Other thrombin-dependent coagulation assays are affected by lepirudin. 

In an APTT clotting assay, which reflects not only the AT III - mediated events but also HC II and other factors of the coagulation cascade besides thrombin and factor Xa, sulfated spaced open chain sugars reached selectivities against... 

The Bethesda assay is used to detect levels of inhibitory antibodies. One Bethesda unit is the reciprocal of the dilution of test plasma at which 50% of normal activity is inhibited. Briefly, the test plasma sample is mixed with an equal volume of pooled normal plasma and incubated at 37 °C for 2 h. Residual activity is measured by aPTT as described above (Herzog et al., 1999 Chao et al., 2001 Fields et al., 2001 Gallo-Penn et al., 2001). 

Specific Factor Assays Factor VIII Factor IX Other Factors Evaluation of the functional concentration (activity) of individual factors, specificity created by using specific factor deficient plasmas. Commonly performed as an aPTT-like assay Sensitivity is to the factor missing from the particular deficient plasma Estimation of the activity level of a particular factor Differences may be observed between deficient plasmas from individuals with hereditary deficiencies and immunodepleted plasmas... 

C4b binding protein can affect the assay The most common assay is a modified aPTT. Elevated levels of Factor VIII bias the assay results, as do decreased levels of vitamin K-dependent factors. 

Specific factor assays are variations on the APTT or PT tests. In the APTT and PT, dilutions of the patient s plasma are made into a deficient, or depleted, substrate plasma. The assays are then performed in the usual way. The clotting times are compared with those obtained from dilutions of pooled normal plasma, commonly 1 10, 1 20, 1 50, and 1 100. A graph of the logarithm of the clotting time (y axis) versus the logarithm of the concentration as percentage of normal x axis) is used to determine the amount of the factor activity in the patient s plasma. The normal pooled plasma is conventionally assigned a value of 100% activity. Many variations exist for specific factor assays, e.g., the venom of Vipera russellii and phospholipids may be substituted for thromboplastin in a PT-like assay. An enzyme in Russell s viper venom rapidly and relatively specifically activates factor X. In conjunction with a factor X-deficient substrate plasma, this provides a specific factor X assay. 

Heparin therapy may be monitored by its increases in clotting time in the APTT, although this method for measuring heparin is difficult to standardize. Heparin is more specifically assayed by its effect on factor Xa inactivation by antithrombin. Such factor Xa-based heparin assays usually employ purified factor Xa as a reagent and factor X-deficient substrate plasma as the source of antithrombin. The prolongation of the clotting time that results from the heparin in the patient s plasma is compared with pooled normal plasma that is known to be free of heparin. Many variations of this heparin assay are available. Heparin assays can use thrombin rather than factor Xa, however, the low-molecular-weight heparins are not reliably measured in thrombin-based assays. 

A young African-American male patient is brought to you because of a hemarthrosis sustained after he twisted his knee while running. His PT is normal, but his APTT is 51 s (normal 20-32 s). Mixing equal volumes of the patient s plasma and normal plasma shortens the APTT to 25 s. Specific factor assays show a normal factor IX activity, but a factor VIII activity of 25% of that in pooled normal plasma. Why was his PT normal What else, prior to the factor IX and VIII assay results, might have been responsible for the prolonged APTT ... 

When used in full therapeutic doses, UFH must be monitored to determine the appropriate dose to administer. The choice of assay is based on chnician preference and institutional availabihty. The aPTT remains the most commonly used test to monitor UFH therapy in North America. In some European countries, anti-factor Xa heparin activity is used commonly. The aPTT should be measured prior to the initiation of therapy to determine the patient s baseline. When administered by intravenous infusion, the response to therapy... 

Accepting the above limitations of our knowledge of the true in vivo reaction sequence, a series of clot-endpoint tests has been used for many years to determine the hemostatic balance in any patient. The first, known as the thrombin time test, can be used to measure the concentration of fibrinogen (B15). The second assay, the activated partial thromboplastin time test (APTT), measures the components of the intrinsic pathway (M4). These include the serine proteases (Factors XII, XI, X, and II), the cofactors (Factors VIII and V) and again, fibrinogen. Most of the hereditary deficiencies... 

In theory, at least, all of the above serine proteases can be assayed by selective specific tests employing synthetic peptide substrates. While these new tests are becoming routine research procedures (S16), the key question for the clinician and laboratory head is how to interpret these results in light of the extensive knowledge of patient testing with the above nonspecific clot-endpoint tests like the PT and APTT. This point will be examined more fully later. 

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