Mixing study (correction test)

Principle

• Patient plasma with a prolonged PT or APTT is mixed 1:1 with normal pooled plasma (NPP, about 100% of each factor).
• A 50% level of any factor is usually enough to give a normal clotting time, so a factor deficiency corrects, while an inhibitor (antibody or drug) also acts on the normal plasma and prevents correction.

Specimen

• 3.2% citrated platelet-poor plasma (double centrifuged if lupus anticoagulant is suspected). Normal pooled plasma must also be platelet poor.

Procedure

• Perform the test that was prolonged (PT or APTT) on patient plasma, NPP and a 1:1 mix immediately.
• For APTT, also incubate patient plasma and NPP separately and a 1:1 mix at 37 °C for 1–2 h; then mix the separately incubated plasmas and test all tubes.

Interpretation

• Immediate and incubated mix correct: factor deficiency; follow with specific factor assays.
• Immediate mix corrects but incubated mix is prolonged: time- and temperature-dependent inhibitor, typical of acquired factor VIII inhibitor.
• Immediate mix does not correct: immediate-acting inhibitor such as lupus anticoagulant, heparin or other anticoagulant drugs.
• Correction criteria vary: into the reference range, or the Rosner index (index of circulating anticoagulant) = (mix time − NPP time) ÷ patient time × 100; a value above about 12–15 suggests an inhibitor (lab validated).

Quality control and pitfalls

• Check for heparin (thrombin time, anti-Xa) before interpreting non-correction.
• Multiple factor deficiencies (e.g. warfarin, liver disease) may correct incompletely.
• Weak inhibitors may appear to correct; strong lupus anticoagulants may make intrinsic factor assays appear low (test at several dilutions).
• NPP should be fresh or properly frozen, with factor levels close to 100%.
• Direct oral anticoagulants cause non-correction.

Clinical use

• Evaluation of unexplained prolonged APTT or PT; distinguishing factor deficiency from lupus anticoagulant or specific factor inhibitors (e.g. in haemophilia patients or acquired haemophilia A).