What should be considered when performing a factor-inhibitor assay?

Updated: Feb 07, 2020
  • Author: Bishnu Prasad Devkota, MD, MHI, FRCS(Edin), FRCS(Glasg), FACP; Chief Editor: Eric B Staros, MD  more...
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If a prolonged PT or aPTT performed on a 1:1 mixture of the patient's plasma and normal plasma leads to suspicion that an inhibitor is present, additional studies can help to determine the inhibitor's nature and titer.

Among immediate-type inhibitors, that is, those that do not require incubation, the existence of heparin in the sample may be the most common cause. This cause can be verified by finding a prolonged thrombin time on a test of the patient's plasma that is corrected with toluidine blue or other agents that neutralize heparin.

Several methods are available for the detection of the lupus anticoagulant, which also requires no incubation. However, lupus anticoagulant is usually associated with a PT that is less prolonged than the aPTT. Moreover, depending on how much phosphatidyl serine exists in each reagent, aPTT reagents differ markedly in their sensitivity to lupus-type anticoagulant. [18]

The development of immunoglobulin inhibitors to specific coagulation factors may occur either (1) after patients with inherited coagulation factor deficiencies have undergone factor replacement therapy or (2) spontaneously in patients with no factor deficiencies. By incubating the patient's plasma with normal plasma, usually for 2 hours at 37°C (98.6°F), and then assaying the specific factor, it is frequently possible to detect antibodies that neutralize factor activity. Although originally designed to quantify factor VIII inhibitors, the Bethesda assay can be modified to detect other coagulation factor inhibitors. [24]

Rather than directly neutralizing clotting activity, some inhibitors instead form complexes with coagulation factors as a means of reducing factor levels, with the complexes rapidly being cleared from the circulation. Such plasmas may be confused with inherited deficiency states because they do not produce prolonged clotting times when mixed 1:1 with normal plasma. Identification of this type of inhibitor requires more elaborate assays; such inhibitors may, for example, cause severe prothrombin deficiency in some patients with antiphospholipid syndrome and deficiency of von Willebrand factor in patients with some acquired forms of von Willebrand disease. [25]

Coagulation factor inhibitors may lead to potentially fatal situations, particularly in patients who are otherwise critically ill. The inhibitor's potency can be measured by titrating the effect of plasma dilutions on the result of mixing the patient's plasma with pooled normal plasma.

Among coagulation factor inhibitors, those directed against factor VIII are the most commonly found. Lupus anticoagulant has been encountered not only in patients with systemic lupus erythematosus (SLE) and drug-induced SLE syndromes, but also in patients with other autoimmune diseases, as well as in otherwise normal persons. A spontaneous reduction in lupus anticoagulants may occur after an offending drug, such as procainamide or a phenothiazine, is discontinued, or may result from the use of immunosuppressive agents such as prednisone.

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