DESCRIPTION

COMP is a secreted pentameric glycoprotein built from modular units and belongs to the thrombospondin protein family. COMP is expressed by hypertrophic chondrocytes and osteoblasts during endochondral ossification, and its expression is increased during fracture healing. COMP is involved in the assembly of collagen fibrils and other structural matrix components. COMP interacts with a number of cartilage ECM proteins, including fibronectin, collagen I, II IX, XII, and XIV, matrilins, as well as proteoglycans such as aggrecan and others.

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	818.1	70.1	8.6
2	20	194.2	18.3	9.4
3	20	51.5	4.6	9.0

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	24	874.3	81.0	9.3
2	24	206.0	20.5	9.9
3	24	44.2	4.4	9.9

RECOVERY

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

Sample Type		Average% of Expected	Range (%)
Human serum	1:160	100	76-127
	1:320	107	87-121
Cell culture supernatant	1:2	105	73-122
	1:4	110	101-122

LINEARITY

To assess the linearity of the assay, cell culture supernatant was spiked with high concentrations of human COMP in various matrices and diluted with the appropriate Sample Diluent to produce samples with values within the dynamic range of the assay. Serum was diluted with the appropriate Sample Diluent to produce samples with values within the dynamic range of the assay.(The serum samples were initially diluted 1:80)

		Human serum	Cell culture supernatant
1:2	Average% of Expected	100	117
	Range (%)	-	115-119
1:4	Average% of Expected	92	121
	Range (%)	83-104	111-130
1:8	Average% of Expected	94	98
	Range (%)	84-106	97-100
1:16	Average% of Expected	94	86
	Range (%)	88-107	85-88

CALIBRATION

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