Myeloperoxidase Assay Kit | MPO assay kit

Human Myeloperoxidase Assay Kit

Cat. Num. AAA480444

• Synonyms: Myeloperoxidase; Human Myeloperoxidase Assay Kit; Human Myeloperoxidase EIA Kit; MPO assay kit

• Sequence Length: 745

• Procedural Notes: 1. Allow kit components to come to room temperature before use.; 2. The HRP conjugate is most stable in its concentrated form. Dilute only the volume necessary for the amount of strips currently being used.; 3. After reconstitution of the MPO Standard, aliquots should be made and frozen at -80 degree C. Avoid multiple freeze/thaw cycles.; 4. To minimize errors in absorbance measurements due to handling, wipe the exterior bottom of the microplate wells with a lint-free paper towel prior to inserting into the plate reader.; 1. The normal ranges for serum samples and heparin-plasma samples are 2.5-100 ng/mL and 37-124; ng/mL, respectively.; 2. Badly hemolyzed serum or plasma samples should not be used, as they may give inaccurate results.; 3. Insufficient washing will result in poor precision and false absorbance readings
• Reagent Preparation: 1. 5x EIA Buffer: Dilute the appropriate amount to 1x with deionized water prior to use.; 2. 20x Wash Buffer: Add 50 mL of 20x Wash Buffer to 950 mL of deionized water prior to use.; 3. Anti-MPO-HRP: Dilute 1:100 - 100 muL of Conjugate into 9.9 mL total volume of 1x EIA; Buffer.; 4. MPO Standard: Reconstitute with 1 mL deionized water.
• Sample Preparation: Thaw samples completely. Dilute the samples in 1x EIA Buffer. 1:4 and 1:10 are recommended
• Standard Curve Preparation: The MPO Standard is provided as a lyophilized powder, and needs to be reconstituted with 1 mL of; deionized water prior to use. Use the following table to construct a six-point standard curve.; Table 1: Standard Curve Preparation; Standard; MPO Conc.; (ng/mL); Vol. of 1x EIA; Buffer (muL); Transfer; Volume (muL); Transfer; Source; Final Volume; (muL); S5 30 538 62 Stock 300; S4 15 300 300 S5 300; S3 7.5 300 300 S4 300; S2 3.75 300 300 S3 300; S1 1.875 300 300 S2 600; B0 0 300 - - 300
• Sample Collection, Storage and Preparation: This ELISA is meant for the measurement of myeloperoxidase in human serum and plasma.; 1. Draw blood into a serum separator tube for serum or a heparin tube for plasma and mix gently.; 2. Centrifuge whole blood within 6 hours of draw at 3000 x g for 10 minutes at 4 degree C.; 3. Remove the serum supernatant. Samples can be stored at 4 degree C for 24 hours. For longer storage, samples should be stored at -20 degree C or -80 degree C. Avoid repeated freeze/thaw cycles.
• Preparation and Storage: 1. Store the components of this kit at the temperatures specified on the labels.; 2. Unopened reagents are stable until the indicated kit expiration date.; 3. Store diluted reagents at 4 degree C and use within one week.; 4. Desiccant packet must remain in the pouch with unused strips. Keep pouch sealed when not in; use to; maintain a dry environment. Remove excess air before sealing.

Related Product Information for MPO assay kit

Introduction: Human myeloperoxidase (MPO) is a hemoprotein composed of two heme-containing heavy subunits of 53 kDa and of two light subunits of 15 kDa. MPO is stored in the granules of neutrophils and monocytes, and is released in response to leukocyte activation. MPO acts as a catalyst in the production of hypochlorous acid (HOCl), a powerful oxidant produced from chloride ion (Cl-) and hydrogen peroxide (H2O2). Although MPO is important to the immune system, it also plays a role in several inflammatory conditions. Elevated MPO levels have been linked to coronary artery diseases. High concentrations of MPO and its oxidation products are found in areas of plaque deposits present in atherosclerosis. MPO also affects cholesterol levels through the oxidation of LDL and HDL. As a specific marker for polymorphonuclear cells (PMNs), MPO is released extracellularly via degranulation after PMN activation. Thus, extracellular MPO is an index of PMN activation in inflammatory processes that lead to disease pathology, such as cardiovascular disease. Elevated levels of MPO present in serum and plasma indicate a greater risk of myocardial injury and unstable plaque formations. Therefore, MPO levels can be used to predict potentially negative cardiac events. Antibodies to MPO are present in human plasma, and have been characterized as part of the antineutrophil cytoplasmic antibodies (ANCA) set of autoantibodies

Principle of assay: The Human MPO EIA Kit is a sandwich ELISA (Enzyme-Linked Immunosorbent Assay) that measures MPO in plasma/serum samples. The MPO in the sample interacts with and binds to the anti-MPO antibody coated on the wells. The plate is then washed to remove all unbound materials and anti-MPO, conjugated with HRP (horseradish peroxidase), is added to the plate wells. The HRP conjugate binds to the MPO that is bound to the anti-MPO of the plate. After washing the plate again, TMB is added, which will produce a blue hue. A darker blue color corresponds to a higher concentration of MPO. After reaction has occurred with the conjugate a stop solution is added, producing a yellow hue. The absorbance is measured at 450 nm.

References

1. Klebanoff, S. J.; (2005) J of Leuk. Bio. 77:598
2. Klein, J.; (1982) Immunology The Science of Self-Nonself Discrimination p 427
3. Renliang, Z., et al.; (2001) JAMA 286(17):2136, 2140
4. Reumaux, D., et al.; (2003) J of Leuk. Bio. 73:846
5. Van der Woode, F.J., and Schmitt, W.H.; (2004) Clinical Applications of Anti-neutrophil Cytoplasmic
Antibody Testing. http://www.medscape.com/viewarticle/467031. Accessed July 31, 2008

NCBI and Uniprot Product Information

• NCBI GI #: 188658
• NCBI GeneID: 4353
• UniProt Accession #: P05164; Q14862; Q4PJH5; Q9UCL7; A1L4B8
• Molecular Weight: 87,248 Da
• NCBI Official Full Name: myeloperoxidase
• NCBI Official Synonym Full Names: myeloperoxidase
• NCBI Official Symbol: MPO
• NCBI Protein Information: myeloperoxidase
• UniProt Protein Name: Myeloperoxidase
• Protein Family: Myeloperoxidase
• UniProt Gene Name: MPO
• UniProt Synonym Gene Names: MPO
• UniProt Entry Name: PERM_HUMAN

NCBI Description

Myeloperoxidase (MPO) is a heme protein synthesized during myeloid differentiation that constitutes the major component of neutrophil azurophilic granules. Produced as a single chain precursor, myeloperoxidase is subsequently cleaved into a light and heavy chain. The mature myeloperoxidase is a tetramer composed of 2 light chains and 2 heavy chains. This enzyme produces hypohalous acids central to the microbicidal activity of neutrophils. [provided by RefSeq, Nov 2014]

Uniprot Description

MPO: Part of the host defense system of polymorphonuclear leukocytes. It is responsible for microbicidal activity against a wide range of organisms. In the stimulated PMN, MPO catalyzes the production of hypohalous acids, primarily hypochlorous acid in physiologic situations, and other toxic intermediates that greatly enhance PMN microbicidal activity. Homodimer; disulfide-linked. Each monomer consists of a light and a heavy chain. Belongs to the peroxidase family. XPO subfamily. 3 isoforms of the human protein are produced by alternative splicing.

Protein type: Oxidoreductase; EC 1.11.2.2

Chromosomal Location of Human Ortholog: 17q23.1

Cellular Component: extracellular space; azurophil granule; mitochondrion; lysosome; nucleus; secretory granule

Molecular Function: heparin binding; peroxidase activity; metal ion binding; heme binding; chromatin binding

Biological Process: respiratory burst during defense response; response to food; removal of superoxide radicals; response to mechanical stimulus; hydrogen peroxide catabolic process; defense response; response to lipopolysaccharide; response to yeast; response to oxidative stress; defense response to fungus; negative regulation of apoptosis; aging

Disease: Myeloperoxidase Deficiency; Alzheimer Disease