DESCRIPTION

MPO(myeloperoxidase) is a peroxidase enzyme presented in the azurophilic granules of polymorphonuclear (PMN) leukocytes and monocytes. Plasma concentration of MPO can be used as a specific marker of PMN activation. MPO catalyzes the production of hypochlorous acid (HClO) from hydrogen peroxide (H2O2) and chloride anion (Cl-, or the equivalent from a non-chlorine halide). This enzymatic system plays an important role in human defense against microorganisms. The serum/plasma MPO levels have been associated with a variety of clinical conditions including inflammatory diseases, atherosclerosis, ischaemic stroke, hypertension, heart failure, risk of cardiovascular events and so on.

PRINCIPLE OF THE ASSAY

An anti-tag antibody is pre-coated onto the bottom of wells. After adding antigen or samples, Tagged-Capture antibody and HRP-conjugated detection antibody, a sandwich complex is formed in the solution. TMB acts as a HRP substrate, and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns the solution yellow. The color intensity is proportional to the quantity of bound protein, which is measurable at 450 nm with the correction wavelength set at 630 nm.

PRECISION

Intra-assay Precision (Precision within an assay) Three samples of known concentration were tested 8 times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in 16 separate assays to assess inter-assay precision.

Intra-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	8	1,242.7	31.5	2.5
2	8	292.6	7.5	2.6
3	8	143.4	5.1	3.5

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	1,259.0	34.6	2.8
2	16	307.2	16.8	5.5
3	16	149.2	8.5	5.7

RECOVERY

The recovery of human MPO spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type		Average% of Expected	Range (%)
Human serum	1:3,200	108	101-121
Cell culture supernatant	1:2,560	105	91-118
Urine	1:16	109	108-110
Saliva	1:6,400	90	88-95
Cell lysate	1:16	110	100-127

LINEARITY

To assess the linearity of the assay, samples were diluted with the appropriate Sample Diluent to produce samples with values within the dynamic range of the assay. (The human serum was initially diluted 1:200. The cell culture supernatant was initially diluted 1:160. The saliva was initially diluted 1:400. The cell lysate was initially diluted 1:10,000.)

		Human serum	Cell culturesupernatant	Urine	Saliva	Cell lysate
1:2	Average% ofExpected	100	100	100	100	100
	Range (%)	-	-	-	-	-
1:4	Average% ofExpected	88	85	91	88	95
	Range (%)	81-95	82-87	89-94	78-98	84-105
1:8	Average% ofExpected	120	93	92	82	81
	Range (%)	111-130	88-97	77-108	77-87	80-82
1:16	Average% ofExpected	106	97	102	84	89
	Range (%)	95-116	91-103	88-116	70-98	79-99

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human MPO produced at Sunlong Medical™.