Mouse CX3CL1/Fractalkine Quantikine ELISA Kit Summary

Product Summary

The Quantikine Mouse Fractalkine Immunoassay is a 4.5 hour solid-phase ELISA designed to measure mouse Fractalkine in cell culture supernates, tissue homogenates, mouse serum, and plasma. It contains E. coli-expressed recombinant mouse Fractalkine chemokine domain and antibodies raised against the recombinant protein. Results obtained for naturally occurring soluble mouse Fractalkine containing the chemokine domain and the mucin stalk domain 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 natural mouse Fractalkine.

Preparation and Storage

Background: CX3CL1/Fractalkine

CX3CL1/Fractalkine is a transmembrane adhesion protein with a chemokine domain separated from the membrane by a mucin stalk. It is upregulated on many cell types during inflammation. The chemokine and mucin regions can be shed as a soluble chemokine that signals through the receptor CX3CR1. During extravasation, membrane-bound CX3CL1 traps leukocytes, then is cleaved to allow diapedesis. Soluble CX3CL1 protects neurons from microglial neurotoxicity, recruits macrophages for wound healing, recruits osteoclast precursors for bone resorption, and contributes to pathogenesis in coronary artery disease.

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.