Hemosil liquid heparin kit

We measured anti-factor Xa activity by performing a chromogenic assay by using the HemosIL Liquid Heparin kit [6] (link), [7] (link) (Instrumentation Laboratory), PT (RecombiPlasTin; HemosIL, Instrumentation Laboratory), and aPTT (SynthASil; HemosIL, Instrumentation Laboratory) in 188 blood samples from the 70 subjects. The blood samples were collected in 3.2% sodium citrate tubes. Anti-Xa activity, PT, and APTT were measured by using the analyzer system ACL TOP 500 CTS (Instrumentation Laboratory). The principle of the anti-factor Xa assay is shown in Fig. 1. The anti-factor Xa activity measurement was expressed as heparin activity (IU/mL). The plasma concentration of rivaroxaban was also estimated by using the following formula: 1 IU/mL=225 ng/mL [10] .

The HemosIL Liquid Heparin Kit (Instrumentation Laboratory, Lexington, KY, USA) was used for the chromogenic AXA assay. This single assay is based on the reaction between FXa and a synthetic chromogenic substrate. For liquid heparin, 10 µL of plasma was mixed with 100 µL of chromogenic substrate and incubated at 37 °C; thereafter, 75 µL of bovine FXa was added. The chromogenic substrate was hydrolyzed by FXa to release p-nitroaniline, and the quantity of p-nitroaniline released was determined using spectrophotometry at 405 nm on an ACL TOP 500 CTS analyzer system (Instrumentation Laboratory). Standard HemosIL heparin calibrators (Instrumentation Laboratory) were used, and AXA was expressed in heparin international units (IU)/mL. To ensure measurement at precise intervals corresponding to the trough concentration, the first blood collection was performed immediately before edoxaban intake in the morning. A second blood collection was performed 2 h later to accurately measure the peak concentration. This timing was based on reaching a peak plasma concentration in 1–2 h of taking edoxaban [11 (link), 17 (link)]. To measure the steady state, serum creatinine level at trough and AXA concentrations at trough and peak times after repeated edoxaban intake were measured at least 72 h after the start of treatment.