Investigation of an anti-activated factor X (anti-Xa) assay for the quantification of enoxaparin in human plasma

• Published in: Journal of pharmacy and pharmacology 2015-02, Vol.67 (2), p.209-214
• Main Authors: Al-Sallami, Hesham S., Medlicott, Natalie J.
• Format: Article
• Language: English
• Subjects: anti-Xa; Anticoagulants - blood; Anticoagulants - therapeutic use; Antithrombin III - metabolism; chromogenic; dalteparin; enoxaparin; Enoxaparin - blood; Enoxaparin - therapeutic use; Factor Xa - metabolism; Freezing; haemolysis; Hemolysis; Humans; Male; Thrombosis - drug therapy
• ISSN: 0022-3573; 2042-7158
• DOI: 10.1111/jphp.12333

Objectives Enoxaparin is a low‐molecular weight heparin (LMWH) widely used for the treatment of thrombosis and measured through a chromogenic assay (anti‐Xa). The aim of this project was to investigate the impact of patient and sampling sources of variation on an anti‐Xa assay for enoxaparin. Methods COATEST was used in accordance with manufacturer's instructions. A standard curve of LMWH concentration (0.1–1.0 IU/ml) was prepared. The shelf‐life and freeze‐thaw stability of Xa over a 6‐month period were investigated. The effects of blood sample haemolysis and plasma antithrombin‐III (AT) concentration were examined. Key findings The standard curve performed well with high accuracy (average bias of 8.1%) and precision (average CV of 2.8%). The shelf‐life of Xa once reconstituted could be extended from 1 month to 4 months if aliquots were frozen at −20°C. Some loss of anti‐Xa activity was observed on freeze‐thawing (bias up to 25%). Haemolysis of 2.5% was found to interfere with the assay. AT level as low as 50% did not affect assay accuracy. Conclusions Xa standards when frozen immediately after reconstitution could be used for the following 4 months. Haemolysis interferes with the assay, but a 50% reduction in AT does not significantly affect the assay result.