Mouse BAFF/BLyS/TNFSF13B Quantikine ELISA Kit Summary

Product Summary

The Quantikine Mouse BAFF/BLyS Immunoassay is a 4.5 hour solid-phase ELISA designed to measure mouse BAFF/BLyS in cell culture supernates, serum, and plasma. It contains NS0-expressed recombinant mouse BAFF/BLyS and antibodies raised against the recombinant factor. This immunoassay has been shown to accurately quantitate the recombinant factor. Results obtained using natural mouse BAFF/BLyS 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 mouse BAFF/BLyS.

Preparation and Storage

Background: BAFF/BLyS/TNFSF13B

BAFF (B-cell activating factor), also known as BLyS, TALL-1, THANK, and TNFSF13B, is a TNF superfamily ligand that plays a critical role in the development and survival of B lineage cells. It can be expressed as a homotrimer or as a heteromer in association with the related protein APRIL. It is produced by many hematopoietic cell types, and a soluble form can be released by activated neutrophils. Both BAFF and APRIL signal through the receptors BCMA and TACI, and BAFF additionally signals through BAFF R. Mice that overexpress BAFF exhibit elevated B cell numbers, increased formation and size of germinal centers, and symptoms of autoimmunity. In addition, BAFF costimulates T cell activation, promotes a Th1 biased immune response, and promotes the expansion of Treg cells.

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.