DESCRIPTION

CXCL5 is a member of the ELR+ CXC chemokine family, whose members contain a highly conserved three amino acid motif (ELR+) that promotes angiogenesis and is highly associated with aberrant angiogenesis. CXCL5 is an important chemokine secreted by immune cells, such as monocytes and T lymphocytes. Recent studies indicated that CXCL5 is aberrantly expressed in >14 different types of cancer, including hepatocellular carcinoma, prostate cancer, pancreatic cancer, and gastric cancer. Antibody neutralization of CXCL5 in experimental models of human non-small-cell lung cancer decreased tumor angiogenesis and metastasis.

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	110.7	8.3	7.5
2	8	27.3	1.7	6.2
3	8	13.9	0.9	6.5

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	109.6	7.8	7.1
2	16	27.2	2.1	7.7
3	16	13.9	0.6	4.3

RECOVERY

The recovery of human CXCL5 spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type		Average% of Expected	Range (%)
Human serum	1:16	110	100-122
	1:32	102	98-108
Cell culture supernatant	1:40	89	87-91
	1:80	90	88-93

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 1:2. The cell culture supernatant was initially diluted 1:20.)

		Human serum	Cell culture supernatant
1:2	Average% of Expected	100	100
	Range (%)	-	-
1:4	Average% of Expected	91	88
	Range (%)	89-92	87-88
1:8	Average% of Expected	84	83
	Range (%)	83-85	82-84
1:16	Average% of Expected	83	79
	Range (%)	82-84	77-82

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human CXCL5 produced at Sunlong Medical™.