A fluorescent homogeneous assay for myeloperoxidase measurement in biological samples. A positive correlation between myeloperoxidase-generated HOCl level and oxidative status in STZ-diabetic rats

• Published in: Talanta (Oxford) Vol. 170; pp. 119 - 127
• Main Authors: Stocker, Pierre, Cassien, Mathieu, Vidal, Nicolas, Thétiot-Laurent, Sophie, Pietri, Sylvia
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01-08-2017
• Subjects: 7-hydroxy-2-oxo-2H-chromene-8-carbaldehyde oxime; Animals; Diabetes Mellitus, Experimental - metabolism; Enzyme Assays - methods; Fluorimetric assay; Halogenation; HL-60 Cells; Humans; Hypochlorous acid; Hypochlorous Acid - metabolism; Male; Myeloperoxidase; Oxidative Stress; Peroxidase - blood; Peroxidase - metabolism; Rats; Spectrometry, Fluorescence; GSSG; 7-hydroxy-2-oxo-2H-chromene-8-carbaldehyde oxime; PC; Hypochlorous acid; Fluorimetric assay; STZ; Myeloperoxidase; 7-HCCO; MPO; GSH
• ISSN: 0039-9140; 1873-3573
• DOI: 10.1016/j.talanta.2017.03.102

Myeloperoxidase (MPO) is a key enzyme derived from leukocytes which is associated with the initiation and progression of many inflammatory diseases. Increased levels of MPO may contribute to cellular dysfunction and tissues injury by producing highly reactive oxidants such as hypochlorous acid (HOCl). Myeloperoxidase-generated HOCl is therefore considered as a relevant biomarker of oxidative stress-related damage and its quantitation is of great importance to the study of disease progression.
In this context, the current study describes a rapid, sensitive and homogeneous fluorescence-based method for detecting the MPO chlorination activity in biological samples. This assay utilizes 7-hydroxy-2-oxo-2H-chromene-8-carbaldehyde oxime as a selective probe for HOCl detection, and is adapted to 96-well microplates to allow high-throughput quantitation of active MPO.
The ability of the method to monitor HOCl release was further investigated in hyperglycemic streptozotocin-treated diabetic rats. The data proved that the present assay has a reliable performance when quantitating the active MPO in the plasma of diabetic animals, a feature of inflammatory disease found concomitant with an elevation of protein carbonyls levels and lipid peroxidation and which was negatively correlated with the ratio of reduced-to-oxidized glutathione.
The method for detecting the MPO chlorination activity utilizes 7-hydroxy-2-oxo-2H-chromene-8-carbaldehyde oxime as a selective probe for HOCl detection.
This assay is adapted to 96-well microplates to allow high-throughput quantitation of active MPO in biological samples. [Display omitted]
•A sensitive and homogeneous method for detecting the MPO chlorination activity.•Evaluation of active MPO level in diabetic animals.•A positive correlation between MPO level and oxidative status in diabetic rats.