DESCRIPTION

S100A8 and S100A9 (also known as MRP8 and MRP14, respectively) are Ca2+ binding proteins belonging to the S100 family. Calprotectin, is a heterodimeric calcium-binding protein consisting of S100A8 and S100A9 subunits. They are expressed in a wide variety of cell types and are abundant in myeloid cells such as: neutrophils, monocytes, keratinocytes and early differentiation states of macrophages. S100A8 and S100A9 have already been confirmed to play a decisive role in the development of inflammation. Elevated levels of S100A8/S100A9 were detected in inflammation, neoplastic tumor cells and various human cancers.

PRINCIPLE OF THE ASSAY

Sandwich ELISA structure (Detection antibody labeled with biotin) A capture antibody is pre-coated onto the bottom of wells which binds to analyte of interest. A detection antibody labeled with biotin also binds to the analyte. Streptavidin-HRP binds to the biotin. TMB acts as the HRP substrate and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns 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 20 times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in 24 separate assays to assess inter-assay precision.

Intra-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	20	1,816.5	76.8	4.2
2	20	413.2	18.6	4.5
3	20	92.4	6.1	6.6

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	24	1,994.1	126.3	6.3
2	24	457.5	38.9	8.5
3	24	115.4	11.1	9.7

RECOVERY

The recovery of human S100A8/S100A9 spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type		Average% of Expected	Range (%)
Human serum	1:10,000	92	79-123
	1:20,000	92	85-102
Cell culture supernatant	1:1,000	93	81-106
	1:2,000	91	87-100
Human milk	1:400	89	76-122
	1:800	89	81-101
Urine	1:5	83	76-96
	1:10	77	76-81
Saliva	1:10,000	98	84-116
	1:20,000	89	78-119

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 samples and saliva were initially diluted 1:500, cell culture supernatant was initially diluted 1:25, the human milk were initially diluted 1:100, the urine were initially diluted 1:2.5)

		Human serum	Cell culturesupernatant	Human milk	Urine	Saliva
1:2	Average% ofExpected	100	100	100	100	100
	Range (%)	-	-	-	-	-
1:4	Average% ofExpected	88	102	90	97	93
	Range (%)	86-92	95-107	87-94	94-103	92-95
1:8	Average% ofExpected	105	89	88	103	95
	Range (%)	82-122	81-104	86-89	96-117	82-106
1:16	Average% ofExpected	79	88	89	97	102
	Range (%)	76-82	83-92	88-90	90-103	85-111

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human S100A8/S100A9 Sandwich ELISA Kit produced at Sunlong Medical™.

Ordering & shipping

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