Human Soluble gp130 Quantikine ELISA Kit Summary

Product Summary

The Quantikine Human Chemerin Immunoassay is a 3.5 hour solid-phase ELISA designed to measure human Chemerin in cell culture supernates, serum, and plasma. It contains E. coli-expressed recombinant human Chemerin and has been shown to accurately quantitate the recombinant factor. Results obtained using natural human Chemerin showed linear curves that were parallel to the standard curves obtained using the Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for naturally occurring human Chemerin.

Preparation and Storage

Background: Chemerin

Chemerin, also known as Tazarotene-induced Gene 2, (Tig-2), is a member of the Cystatin superfamily. Chemerin is synthesized as a 163 aa precursor that contains a hydrophobic 20 aa N-terminal sequence, an intervening 137 aa Cystatin fold-containing domain, and a six aa C-terminal prosegment. It is activated by proteases of the coagulation, fibrinolytic, and inflammatory cascades. Chemerin is a chemoattractant for cells bearing the Chem R23 receptor, including plasmacytoid dendritic cells and tissue macrophages.

Assay Procedure

Refer to the product datasheet for the complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.

1.     Prepare all reagents, standard dilutions, and samples as directed in the product insert.

2.     Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

3.     Add 50 μL of Assay Diluent to each well.

4.     Add 50 μL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.

5.     Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.

6.     Add 100 μL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.

7.     Aspirate and wash 5 times.

8.     Add 100 μL Substrate Solution to each well.

9.     Add 100 μL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.