Consider this scenario: inpatient with bruising has tests for coagulation. The APTT is prolonged at 40 s (normal <35). Is this abnormal? Is it due to the prophylactic clexane he is receiving? How do you tell?
Heparin contamination of inpatient samples is common. However, the causes of a prolonged APTT with normal PT are varied, and must be considered in an (usually) elderly patient with ecchymoses and bruises. Aetiologies include heparin, LMWH, low dose dabigatran, inherited deficiencies of factor VIII, IX and XI or VWF in young persons(with bleeding) or factor XII, HMWK or prekallikrein (without bleeding) and ciculating inhibitors to Factor VIII (commonest), seen typically in postpartum women, in autoimmune disease such as SLE and RA, and in cancer, or inhibitors to factor IX or XI.
If heparin contamination is suspected in a repeatedly prolonged APTT (always repeat the test), the best test is a combination of thrombin time and reptilase time. Thrombin time (TT), performed by using the patient's plasma with human or bovine thrombin, will be prolonged in heparin contamination, while reptilase time (RT), performed by mixing the patient's plasma with a venom obtained from Bothrops snake remains normal. The venom, unlike thrombin, cannot be inhibited by heparin or LMWH or by direct thrombin inhibitors.
Alternatively, use an inhibitor of heparin such as protamine or a heparin binding resin such as Heparsorb and repeat the test. APTT will normalise if it was prolonged due to heparin.
In cases of hypofibrinogenemia or dysfibrinogenemia, both TT and RT will be prolonged.
A circulating inhibitor is often suspected in adults with ecchymoses or bruising, occasionally with associated joint swelling due to haemarthrosis, prolonged APTT, normal PT, and no alternative causes such as liver failure, DIC (usually pronlongs both PT & APTT) or concurrent anticoagulant adminstration. The commonest circulating inhibitor is to factor VIII. While this is typically seen in haemophiliacs, it can also be seen in postpartum women, usually primigravidas, and as stated above, with autoimmune disease or cancer.
After ruling out heparin contamination as above, the patient's plasma is incubated 1:1 with normal plasma for 1 to 2 hours at 37 degrees celsius (to allow stable binding of IgG antibodies, which work best at body temperature) . APTT will correct with factor VIII, IX, or XI deficiencies or with vWD. If the APTT does not correct, it's either a circulating inhibitor, usually to Factor VIII, or lupus anticoagulant. The latter presents with thrombosis rather than bleeding and can be easily ruled out by one of two tests. First, adding phospholipid (usually cephalin from brain tissue) normalises the APTT in subjects with lupus anticoagulant. Here, one must be careful in performing a hexagonal phase assay rather than a lamellar assay. Hexagonal phase assay includes adding phosphatidylethanolamine rather than phospatidylcholine, as is found in lamellar (bilipid layer) assays. Although the latter is found in physiological membranes, circulating lupus anticoagulant is only inhibited by phospholipids present in the hexagonal form, comprising packed phospatidylethanolamine in a six-cylinder configuration. Using lipids in a lamellar form will lead to a false negative test.
The second test is to simply repeat the clotting with the dilute Russell Viper Venom Time (dRVVT). This bypasses all factors upstream of Factor X. Since Factor X still needs phospholipids to activate thrombin, it is abnormal with antiphospholipid syndrome, but not with circulating Factor VIII inhibitor.
It is highly likely that a person with prolonged APTT that does not correct with normal plasma, and with a normal dRVVT has a circulating inhibitor to Factor VIII. The next step is the Bethesda assay. Here, increasing dilutions of the patient's plasma is added to normal plasma, and assay is performed for Factor VIII activity, as in haemophilia. The reciprocal of the dilution that produces 50% of the normal Factor VIII activity is expressed in Bethesda Units (BU). More than 5 BU is considered abnormal.
Therefore, I'd suggest the following algorithm when one comes across an unexplained rise in APTT with normal PT.
1. Repeat test.
2. If APTT still prolonged, rule out heparin contamination by checking Thrombin Time and Reptilase Time. If TT is prolonged and RT is normal, heparin is present. If both TT and RT are prolonged, proceed to the next step.
3. Dilute the patient's plasma 1:1 with normal plasma and incubate for 60 minutes at 37 degrees Celsius. If APTT normalises, the patient has deficiency of one of the following factors: Factor VIII, IX, XI, VWF, Factor XII, HMWK, or prekallekrein. Deficiency of one of the last three prolongs the APTT, but does not lead to bleeding.
4. If the APTT does not correct with addition of normal plasma, the patient either has lupus anticoagulant or a circulating inhibitor to one of Factor VIII (commonest), or Factor IX or XI. Lupus anticoagulant leads to clotting rather than bleeding in vivo.
5. Test for lupus anticoagulant by doing one of these two tests: Do the dilute Russell Viper Venom Time. If dRVVT is prolonged, patient has Lupus anticoagulant.
Alternatively, add phospholipid to the mixture of normal and patient plasma and repeat APTT. If the latter returns to normal, lupus anticoagulant is present.
6. If the dRVVT is normal or if APTT is not corrected by adding phospholipid, a circulating inhibitor to Factor VIII, IX or XI is present. Inhibitors to Factor VIII are by far the commonest, and should particularly be suspected in postpartum women, usually after their first child, or older subjects. In such cases, proceed directly to the Bethesda assay. Presence of more than 5 BU is consistent with presence of Factor VIII inhibitor.
If heparin contamination is suspected in a repeatedly prolonged APTT (always repeat the test), the best test is a combination of thrombin time and reptilase time. Thrombin time (TT), performed by using the patient's plasma with human or bovine thrombin, will be prolonged in heparin contamination, while reptilase time (RT), performed by mixing the patient's plasma with a venom obtained from Bothrops snake remains normal. The venom, unlike thrombin, cannot be inhibited by heparin or LMWH or by direct thrombin inhibitors.
Alternatively, use an inhibitor of heparin such as protamine or a heparin binding resin such as Heparsorb and repeat the test. APTT will normalise if it was prolonged due to heparin.
In cases of hypofibrinogenemia or dysfibrinogenemia, both TT and RT will be prolonged.
A circulating inhibitor is often suspected in adults with ecchymoses or bruising, occasionally with associated joint swelling due to haemarthrosis, prolonged APTT, normal PT, and no alternative causes such as liver failure, DIC (usually pronlongs both PT & APTT) or concurrent anticoagulant adminstration. The commonest circulating inhibitor is to factor VIII. While this is typically seen in haemophiliacs, it can also be seen in postpartum women, usually primigravidas, and as stated above, with autoimmune disease or cancer.
After ruling out heparin contamination as above, the patient's plasma is incubated 1:1 with normal plasma for 1 to 2 hours at 37 degrees celsius (to allow stable binding of IgG antibodies, which work best at body temperature) . APTT will correct with factor VIII, IX, or XI deficiencies or with vWD. If the APTT does not correct, it's either a circulating inhibitor, usually to Factor VIII, or lupus anticoagulant. The latter presents with thrombosis rather than bleeding and can be easily ruled out by one of two tests. First, adding phospholipid (usually cephalin from brain tissue) normalises the APTT in subjects with lupus anticoagulant. Here, one must be careful in performing a hexagonal phase assay rather than a lamellar assay. Hexagonal phase assay includes adding phosphatidylethanolamine rather than phospatidylcholine, as is found in lamellar (bilipid layer) assays. Although the latter is found in physiological membranes, circulating lupus anticoagulant is only inhibited by phospholipids present in the hexagonal form, comprising packed phospatidylethanolamine in a six-cylinder configuration. Using lipids in a lamellar form will lead to a false negative test.
The second test is to simply repeat the clotting with the dilute Russell Viper Venom Time (dRVVT). This bypasses all factors upstream of Factor X. Since Factor X still needs phospholipids to activate thrombin, it is abnormal with antiphospholipid syndrome, but not with circulating Factor VIII inhibitor.
It is highly likely that a person with prolonged APTT that does not correct with normal plasma, and with a normal dRVVT has a circulating inhibitor to Factor VIII. The next step is the Bethesda assay. Here, increasing dilutions of the patient's plasma is added to normal plasma, and assay is performed for Factor VIII activity, as in haemophilia. The reciprocal of the dilution that produces 50% of the normal Factor VIII activity is expressed in Bethesda Units (BU). More than 5 BU is considered abnormal.
Therefore, I'd suggest the following algorithm when one comes across an unexplained rise in APTT with normal PT.
1. Repeat test.
2. If APTT still prolonged, rule out heparin contamination by checking Thrombin Time and Reptilase Time. If TT is prolonged and RT is normal, heparin is present. If both TT and RT are prolonged, proceed to the next step.
3. Dilute the patient's plasma 1:1 with normal plasma and incubate for 60 minutes at 37 degrees Celsius. If APTT normalises, the patient has deficiency of one of the following factors: Factor VIII, IX, XI, VWF, Factor XII, HMWK, or prekallekrein. Deficiency of one of the last three prolongs the APTT, but does not lead to bleeding.
4. If the APTT does not correct with addition of normal plasma, the patient either has lupus anticoagulant or a circulating inhibitor to one of Factor VIII (commonest), or Factor IX or XI. Lupus anticoagulant leads to clotting rather than bleeding in vivo.
5. Test for lupus anticoagulant by doing one of these two tests: Do the dilute Russell Viper Venom Time. If dRVVT is prolonged, patient has Lupus anticoagulant.
Alternatively, add phospholipid to the mixture of normal and patient plasma and repeat APTT. If the latter returns to normal, lupus anticoagulant is present.
6. If the dRVVT is normal or if APTT is not corrected by adding phospholipid, a circulating inhibitor to Factor VIII, IX or XI is present. Inhibitors to Factor VIII are by far the commonest, and should particularly be suspected in postpartum women, usually after their first child, or older subjects. In such cases, proceed directly to the Bethesda assay. Presence of more than 5 BU is consistent with presence of Factor VIII inhibitor.
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