Patent EP-1711817-B1 (EP)
Inventor: SERTEYN DIDIER
IPC: G01N33/573
Filed: 2005-02-07
Granted: 2008-11-26
Abstract: In vitro measurement of neutrophil cell activation status in a sample (preferably from horse) comprising obtaining a sample from a horse containing the neutrophil cells and/or MPO released by the cells; (immuno)capturing myeloperoxidase (MPO) present; detecting and/or measuring either total (active and inactive) MPO or exclusively active MPO; and comparing the measured MPO values with normal MPO levels from healthy horses, is new. In vitro measurement of neutrophil cells activation status in a mammalian sample (preferably from horse) by specifically correlating the myeloperoxidase (MPO) content with the cell activation status comprising: (a) obtaining a biological sample (a fluid from the horses) containing the neutrophil cells and/or MPO released by the cells; (b) (immuno)capturing MPO present in the sample via a MPO-specific polyclonal or monoclonal antibody; (c) detecting and/or measuring either total (active and inactive) MPO or exclusively active MPO present in the sample; (d) comparing the measured MPO values with normal MPO levels obtained from healthy horses; (e) optionally quantifying MPO levels using a standard MPO curve; and (f) relating the MPO levels measured to an activity status of the cells indicative of the presence, absence or condition of a disease or immunological status. Independent claims are also included for: (1) an ELISA kit or device (for measuring the activation status of neutrophil cells in a biological sample obtained from a mammal, preferably a horse, by specifically correlating the MPO content with the cell activation status) comprising the necessary elements for immunocapturing MPO in the sample (preferably a horse sample containing the neutrophil cells or MPO released by the cells) being preferably obtained by the first MPO recognizing polyclonal or monoclonal antibody immobilized to a solid support, detecting and/or measuring active and inactive MPO present in the sample, preferably by a second enzymatically labeled MPO-recognizing polyclonal or monoclonal antibody for detection of immunocaptured MPO; possibly comparing the measured MPO values with normal MPO levels obtained from healthy mammals, optionally quantifying MPO levels using a standard MPO curve, and possibly relating the MPO levels measured to an activity status of the cells indicative of the presence, absence or condition of a disease or immunological status, where the detection and/or measurements specifically and accurately representing MPO levels in any type of biological sample; (2) a SIEFED kit or device (for measuring the activation status of neutrophil cells in a biological sample obtained from a mammal, preferably a horse, by specifically correlating the MPO content with the cell activation status) comprising the necessary elements for: immunocapturing MPO in the sample (preferably a biological fluid, from, a horse and containing the neutrophil cells or MPO released by the cells), detecting and/or measuring active MPO present in the biological sample, where nitrite may be added to the reaction medium to amplify a MPO enzymatic reaction, possibly comparing the measured MPO values with normal MPO levels obtained from healthy mammals (preferably horses), optionally quantifying MPO levels using a standard MPO curve, and possibly relating the MPO levels measured to an activity status of the cells indicative of the presence, absence or condition of a disease or immunological status, where the detection and/or measurements specifically and accurately representing active MPO levels in any type of biological sample; (3) using nitrite, preferably its sodium salt, is used to enhance enzymatic detection of the peroxidase, preferably to enhance a 10-acetyl-3,7-dihydroxyphenoxazine induced fluorescence signal; (4) using both an ELISA and a SIEFED method to distinguish between total and active MPO content; and (5) screening and selecting compound(s) (interacting with MPO, preferably inhibiting MPO activity) comprising capturing active MPO to a solid support, measuring MPO activity, measuring MPO activity by adding one or more compound(s) to the active MPO, comparing MPO activities before or after addition of the compounds, and recovering the compound(s) that interact with MPO, preferably compounds that inhibit myeloperoxidase activity.