Human CD25/IL-2R alpha Quantikine ELISA Kit Summary

Product Summary

The Quantikine Human IL-2 sR alpha Immunoassay is a 3.5 hour solid phase ELISA designed to measure IL-2 sR alpha in cell culture supernates, serum, and plasma. It contains NS0-expressed recombinant human IL-2 sR alpha and antibodies raised against the recombinant factor. This immunoassay has been shown to accurately quantitate the recombinant factor. Results obtained using natural human IL-2 sR alpha showed linear curves that were parallel to the standard curves obtained using the recombinant Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for natural human IL-2 sR alpha.

Preparation and Storage

Background: CD25/IL-2R alpha

CD25/IL-2 receptor alpha is a transmembrane protein that associates with IL-2 R beta/CD122 and the Common gamma Chain/CD132 on activated T cells and regulatory T cells (Treg). The Common gamma chain is a shared subunit in the receptors for IL-4, -7, -9, -15, and -21. IL-2 R alpha is required for mediating IL-2 induced effects including T cell proliferation, activation induced cell death (AICD) of autoreactive naive T cells, and the development of CD4+CD25+ Treg which contribute to peripheral T cell homeostasis. A soluble form of IL-2 R alpha can be generated by proteolytic cleavage of the cell surface receptor.

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.