Description

Nori Equine IL-28A ELISA Kit Summary

Alternative names for IL-28A: IL28A, interleukin 28A, interferon lambda 2, IFNL2

Alternative name for equine: Horse

Standard Curve

Kit Components
1. Pre-coated 96-well Microplate
2. Biotinylated Detection Antibody
3. Streptavidin-HRP Conjugate
4. Lyophilized Standards
5. TMB One-Step Substrate
6. Stop Solution
7. 20 x PBS
8. Assay Buffer

Other Materials Required but not Provided:
1. Microplate Reader capable of measuring absorption at 450 nm
2. Log-log graph paper or computer and software for ELISA data analysis
3. Precision pipettes (1-1000 µl)
4. Multi-channel pipettes (300 µl)
5. Distilled or deionized water

Protocol Outline
1. Prepare all reagents, samples and standards as instructed in the datasheet.
2. Add 100 µl of Standard or samples to each well and incubate 1 h at RT.
3. Add 100 µl of Working Detection Antibody to each well and incubate 1 h at RT.
4. Add 100 µl of Working Streptavidin-HRP to each well and incubate 20 min at RT.
5. Add 100 µl of Substrate to each well and incubate 5-30 min at RT.
6. Add 50 µl of Stop Solution to each well and read at 450 nm immediately.

Background: 

Interleukin-28 (IL-28) is a cytokine that comes in two isoforms, IL-28A and IL-28B, and plays a role in immune defense against viruses, including the induction of an “antiviral state” by turning on Mx proteins, 2′,5′-oligoadenylate synthetase as well as ISGF3G (Interferon Stimulated Gene Factor 3).^[1] IL-28 has also been shown to play a role in the adaptive immune response, as its inclusion as an immunoadjuvant during small animal vaccination lead to augmented antigen-specific Interferon Gamma release as well as an increased cytotoxic potential in CD8+ T cells.^[2] IL-28A and IL-28B belong to the type III interferon family of cytokines and are highly similar (in amino acid sequence) to IL-29. IL28A, IL28B, and IL29, also named interferon λ2 (IFNλ2), IFNλ3, and IFNλ1, respectively, are class II cytokine receptor ligands that are distantly related to members of the IL10 family (11-13% aa sequence identity) and the type I IFN family (15-19% aa sequence identity). ^[3-5]  The expression of IL-28A, IL-28B, and IL29 is induced by virus infection or double stranded RNA. All three cytokines exert bioactivities that overlap those of type I IFNs, including antiviral activity and upregulation of MHC class I antigen expression. The three proteins signal through the same heterodimeric receptor complex that is composed of the IL10 receptor β (IL10 Rβ) and a novel IL28 receptor α (IL28Rα, also known as IFNλ R1). Ligand binding to the receptor complex induces Jak kinase activation and STAT1 and STAT2 tyrosine phosphorylation. The phosphorylated STAT1 and STAT2 complex with IFN regulatory factor 9 (IRF9) to form the IFN stimulated regulatory factor 3 (ISGF3) transcription factor complex that is translocated to the nucleus. ISGF3 binds to the IFN stimulated response element (ISRE) present in the regulatory region of the target genes.

References

1. Kempuraj D, et al. (2004). Int J Immunopathol Pharmacol. 17(2): 103–6.
2. Morrow MP, et al. (2009).  Blood. 113(23): 5868–77.
3. Vilcek, J. (2003) Nature Immunol. 4:8.
4. Sheppard, P. et al. (2003) Nature Immunol. 4:63.
5. Kotenko, S.V. et al. (2003) Nature Immunol. 4:69.

Product Citations