DESCRIPTION

The chemokine stroma-derived factor (SDF-1/CXCL12) plays multiple roles in tumor pathogenesis. CXCL12 plays an important role in angiogenesis by recruiting endothelial progenitor cells (EPCs) from the bone marrow through a CXCR4 dependent mechanism (PMID: 17878755). It is this function of CXCL12 that makes it a very important factor in carcinogenesis and the neovascularisation linked to tumour progression (PMID: 16943240). CXCL12 also has a role in tumor metastasis where cancer cells that express the receptor CXCR4 are attracted to metastasis target tissues that release the ligand, CXCL12 (PMID: 11242036 ). In breast cancer, however, increased expression of CXCL12 determines a reduced risk of distant metastasis (PMID: 19646861 ).

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	1,795.0	43.0	2.4
2	20	423.5	11.7	2.8
3	20	93.5	3.8	4.1

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	24	2,082.6	100.2	4.8
2	24	462.3	20.1	4.4
3	24	108.7	4.5	4.1

RECOVERY

The recovery of mouse CXCL12/SDF-1 spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type		Average% of Expected	Range (%)
Mouse serum	1:15	94	79-109
	1:30	94	73-126

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.

		Mouse serum
1:15	Average% of Expected	100
	Range (%)	-
1:30	Average% of Expected	107
	Range (%)	92-122
1:60	Average% of Expected	100
	Range (%)	78-123

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Mouse CXCL12/SDF-1 Sandwich ELISA Kit produced at Sunlong Medical™.