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Inhibitor Index in the Clot Waveform Analysis-Based Mixing Test Differentiates among Hemophilia A without and with Inhibitors, and Lupus AnticoagulantFunding This work was partly supported by a Grant-in-Aid for Scientific Research (KAKENHI) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) to K.N. (18K07885).
Background The mixing test is used to identify the pathway to follow-up testing and is also useful for the investigation of lupus anticoagulant (LA) positivity. “To completely correct” indicates clotting factor deficiency, while “to not correct” indicates the presence of a clotting factor inhibitor including LA. “Index of circulation anticoagulant” and/or “percent correction” is used to interpret the results of mixing studies, but it does not accurately differentiate factor inhibitors from LA.
Aim To precisely differentiate hemophilia A (HA), HA with inhibitor (HA-inh), and LA using the clot waveform analysis (CWA)-based mixing test.
Methods Plasma samples from HA, LA, and HA-inh including acquired HA were incubated with normal plasma in 9:1, 1:1, and 1:9 mix ratios. From activated partial thromboplastin time CWA at 0-minute (immediately) and 12-minute incubation, the ratios of CWA parameters at 12 minutes/0 minute (inhibitor index) were assessed.
Results The inhibitor index values of CWA parameters obtained using the mixing test in a 1:1 ratio demonstrated a significant difference between HA-inh and LA but could not differentiate LA from HA-inh completely. Plasmas used for the mixing tests in 9:1 and 1:9 ratios were able to fully distinguish between HA-inh (>0.5 BU/mL) and LA. These indices significantly correlated with inhibitor titer below 40 BU/mL (r > 0.90), possibly estimating FVIII inhibitor titer from the inhibitor index. Plasmas in HA and LA could be distinguished by mixing in a 1:1 ratio at 0 minute (immediately).
Conclusion The inhibitor index from CWA-based mixing tests with a 12-minute incubation could differentiate among HA, HA-inh, and LA quickly.
An account of this work was presented at the XXVIII Congress of the International Society on Thrombosis and Haemostasis, 2020, Milano, Italy.
N.S. performed the experiments, analyzed the data, and created the figures. K.O. analyzed and interpreted the data. Y.O., T.K., and M.E. performed the experiments and analyzed and interpreted the data. M.S. supervised this study. K.N. designed all the experiments, interpreted the data, created the figures, wrote and edited the manuscript, and approved the submission of the first version of the manuscript.
Received: 15 August 2020
Accepted: 08 November 2020
07 January 2021 (online)
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