DESCRIPTION

COL6A3 belongs to the type VI collagen family. Collagen VI acts as a cell-binding protein. It is expected to be expressed in smooth muscle. And the calculated molecular weight of COL6A3 is 343 kDa. The protein has similar expression in obese and T2DM (2 patients, and it regulates the chemotaxis and inflammation of macrophages in adipose tissue. Its expression is also related to weight gain. The expression of this protein in adipocytes is related to insulin resistance, which is believed to depend on PPAR (peroxisome proliferator-activated receptor) α-mediated adipocyte development (PMID: 25337653).

PRINCIPLE OF THE ASSAY

Sandwich ELISA structure (Detection antibody labeled with HRP) A capture antibody is pre-coated onto the bottom of wells which binds to analyte of interest. A detection antibody labeled with HRP also binds to the analyte. 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	1,349.5	54.0	4.0
2	20	300.3	7.7	2.6
3	20	85.0	3.1	3.6

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	24	1,386.1	55.4	4.0
2	24	301.9	15.1	5.0
3	24	78.7	3.8	4.9

RECOVERY

The recovery of human COL6A3 spiked to three different levels throughout the range of the assay in human serum was evaluated.

Sample Type		Average% of Expected	Range (%)
Human serum	1:16	98	89-114
	1:32	93	86-102

LINEARITY

To assess the linearity of the assay, human serum 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 1:2.)

		Human serum
1:2	Average% of Expected	100
	Range (%)	-
1:4	Average% of Expected	113
	Range (%)	110-118
1:8	Average% of Expected	93
	Range (%)	86-98
1:16	Average% of Expected	104
	Range (%)	95-117

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human COL6A3 produced at Sunlong Medical™.