DESCRIPTION

Granulysin (GNLY) is a member of the saposin-like protein (SAPLIP) family that is a cytolytic and proinflammatory molecule expressed by activated human cytotoxic T cells and NK cells, exhibiting lytic activity against a variety of microorganisms and tumors. Granulysin is produced as an intact 15-kDa form, and portions are then cleaved at the amino and carboxy termini to produce a 9-kDa form. Granulysin has broad clinical relevance to a wide variety of diseases, including infections, cancer, transplantation, skin afflictions and reproductive complications. Elevation of Granulysin expression and levels in tissue and serum has been reported in infections, autoimmune diseases, transplant rejection. Granulysin expression has been widely correlated with good outcomes in a variety of cancers.

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	1,443.8	25.8	1.8
2	8	332.6	5.7	1.7
3	8	177.6	3.4	1.9

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	1,418.5	33.5	2.4
2	16	329.0	6.3	1.9
3	16	174.7	4.7	2.7

RECOVERY

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

Sample Type		Average% of Expected	Range (%)
Human serum	1:8	114	108-122
	1:16	124	121-125
Cell culture supernatant	1:4	113	105-119

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 cell culture supernatant was initially diluted 1:8.)

		Human serum	Cell culture supernatant
1:2	Average% of Expected	100	100
	Range (%)	-	-
1:4	Average% of Expected	108	106
	Range (%)	93-122	101-112
1:8	Average% of Expected	113	112
	Range (%)	97-128	101-123
1:16	Average% of Expected	115	109
	Range (%)	102-128	107-111

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human GNLY/Granulysin One-Step ELISA Kit produced at Sunlong Medical™.