Fluorescence artifact correction in the thrombin generation assay: Necessity for correction algorithms in...

Area of Science:

• Hemostasis and Thrombosis Research
• Biochemical Assays and Diagnostics
• Clinical Pathology

Background:

• Thrombin generation (TG) assays measure blood clotting but can underestimate hypercoagulability.
• Substrate depletion and inner filter effects (IFE) can cause inaccuracies in TG assays.

Purpose of the Study:

• To determine the necessity of correcting for substrate depletion and IFE in TG assays.
• To evaluate the impact of these corrections on TG parameters under various procoagulant conditions.

Main Methods:

• Analysis of TG parameters using a calibrated automated thrombogram (CAT) platform and in-house software.
• Evaluation in plasma with elevated coagulation factors (I, V, VIII, IX, X, XI) or prothrombin, with/without thrombomodulin.
• Assessment of artifact correction for substrate depletion and IFE.

Main Results:

• Elevated thrombin peak height (TPH) and endogenous thrombin potential (ETP) were observed with increased coagulation factors.
• The CAT algorithm's correction was minimal (<10%) for most elevated factors, not affecting procoagulant sample detection.
• For elevated prothrombin, uncorrected TG values were underestimated, while CAT correction significantly increased TG curves.

Conclusions:

• Correction for substrate consumption and IFE is crucial for identifying specific hypercoagulable states, particularly elevated prothrombin.
• These corrections are not universally required for all procoagulant conditions, such as elevated factors XI and VIII.