Mouse/Rat TNF RII/TNFRSF1B Quantikine ELISA Kit Summary

Product Summary

The Quantikine Mouse/Rat TNF RII Immunoassay is a 4.5 hour solid phase ELISA designed to measure mouse or rat TNF RII in cell culture supernates, serum, and plasma. It contains E. coli-expressed recombinant mouse TNF RII and antibodies raised against the recombinant factor. This immunoassay has been shown to quantitate the recombinant protein accurately. Results obtained using natural mouse or rat TNF RII 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 or rat TNF RII.

Preparation and Storage

Background: TNF RII/TNFRSF1B

TNF RII (Tumor Necrosis Factor Receptor II), also known as TNFRSF1B, p75/p80, and CD120b, is a widely expressed receptor for membrane-associated TNF-alpha and Lymphotoxin-alpha. Its activation initiates pro-inflammatory and pro-survival responses via NFkB-dependent signaling pathways, although it may also induce apoptosis.

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.