Materials Provided

• Pre-coated microtiter plate(s)
• Plate Sealers
• Wash Solution Concentrate
• Mouse Interferon Alpha (IFN-Alpha) Standard, 10,000 pg/ml
• Sample Buffer
• Concentrate Diluent
• Antibody Concentrate
• HRP Conjugate Concentrate
• Assay Diluent
• TMB Substrate Solution
• Stop Solution

Additional Materials Required (Not Provided) 

• Microplate reader capable of reading an OD at a wavelength of 450 nm
• Variable volume microtiter pipettes
• Adjustable multichannel pipette (50-300 μl)
• Reagent reservoirs
• Wash bottle or plate washing system
• Distilled or deionized water
• Serological pipettes (1, 5, 10 or 25 ml)
• Disposable pipette tips (polypropylene)
• Plate shaker

Application Note:

• Differential Biological Activities of E.coli Produced Murine IFN-α Proteins on B16-F10 Cells

Technical Note (by request)

• VeriKine Mouse Interferon Alpha ELISA Kit Use with Tissue Homogenates

Background

Interferons (IFNs) are a group of cytokines which exhibit pleitropic activities that play major roles in both innate and adaptive immunity. Type I IFNs consist of multiple IFN-α genes and at least one IFN-β gene in most vertebrates, and a few other family members such as limitin in the mouse.¹ There are 14 IFN-α subtypes in the mouse which share at least 75% identity in the protein sequence. In the human, IFN-α is used to treat viral diseases and cancer^2-4 and is studied in mouse models of these diseases.^5,6 Recent evidence has suggested that IFN-α may play a role in the development of certain autoimmune diseases^7,8 and that treatment with IFN-α is useful for other autoimmune diseases.⁹

The expression of interferons is regulated by a group of interferon regulatory factor proteins (IRF). The expression of the individual IFN-α subtypes is regulated by the cellular levels of IRF3 and IRF7. Most of the IFN-α subtypes require IRF7 for expression. IRF7 is constitutively expressed in some cell types and is inducible in others. Inducible IRF7 expression is induced by IFN-β and IFN-α4 in mice and leads to expression of the other IFN-α subtypes. Murine IFN-β may be required for the production of IFN-α by fibroblasts but may not be required for IFN-α production by other cells.¹⁰ The IRF3/IRF7 signaling cascade is important for the initial and progressive responses to pathogens wherein hundreds of genes are regulated in a coordinated, temporal manner.¹¹

Plasmacytoid dendritic cells are the major producers of IFN-α, but a large variety of cells can also produce lower levels of these proteins.^12,13 The particular subtypes that are expressed appear to be somewhat cell and stimulus specific.^14-16

104 Citations:

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17. Staehelin, T., Stähli, C., Hobbs, D.S., and Pestka, S. (1981) "A Rapid Quantitative Assay of High Sensitivity for Human Leukocyte Interferon with Monoclonal Antibodies," in Methods in Enzymology, Vol. 79 (S. Pestka, ed.), Academic Press, New York, 589-595.


18. Kelder, B., Rashidbaigi, A., and Pestka, S. (1986) "A Sandwich Radioimmunoassay for Human IFN‑γ gamma," Methods in Enzymology, Vol. 119 (S. Pestka, ed.), Academic Press, New York, 582-587.


19. Mouse IFN-α international reference standard provided by the NIH, reference no. Gxa02-901-511. Pestka, S. (1986) "Interferon Standards and General Abbreviations," Methods in Enzymology, Vol. 119 (S. Pestka, ed.), Academic Press, New York, 14-23.