The VeriKine-HS Human Interferon Alpha All Subtype TCM ELISA Kit is specifically formulated to detect all human IFN-Alpha subtypes. This kit quantitates all human IFN alpha subtypes in tissue culture media (TCM) using a sandwich immunoassay format. The kit is based on an ELISA with biotinylated-detection antibody and streptavidin-conjugated horseradish peroxidase (HRP). Tetramethyl-benzidine (TMB) is the substrate. The assay is based on the international reference standard for human interferon alpha provided by the National Institutes of Health.

*For complex biological matrices such as plasma, healthy sera, and autoimmune disease sera, we recommend our Human IFN-Alpha All Subtype ELISA Kit, High Sensitivity (Cat. No. 41115-1).

Materials Provided

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

Additional Materials Required (Not Provided)

• Microtiter plate reader capable of reading a wavelength of 450 nm
• Plate shaker (recommended shaking capability of 550 rpm)
• Variable volume microtiter pipettes
• Adjustable multi-channel 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)

Tips, Tools and Troubleshooting:

• Human IFN-Alpha ELISA Subtype Specificities Comparison Chart

Representative Standard Curves of Human IFN-Alpha Subtypes

Linearity

Parallelism of Endogenous Samples

sIFNAR2 Receptor Interference

Endogenous Levels of IFN-Alpha Quantified in Sendai Virus (A549)

Endogenous Levels of IFN-Alpha Quantified in Sendai Virus (U937)

Background

Interferons (IFNs) are a family of mammalian cytokines initially characterized by their ability to inhibit viral infection. They are synthesized and secreted by most cell types in response to pathogens. In addition to their antiviral properties, IFNs have also been shown to exhibit anti-proliferative, immunomodulatory, and many other activities.

In humans, IFN-α refers to a family of proteins comprised of 12 highly homologous protein subtypes (greater than 85% by amino acid sequence), encoded by 13 genes, that exhibit pleiotropic biologic activities. Measuring the levels of the 12 different human IFN-α subtypes is essential for understanding their triggers and subsequent effects on the immune system. The ability to measure all the subtypes may provide an indication of the total IFN-α level.

References:

1. Staehelin et al. (1981) "A Rapid Quantitative Assay of High Sensitivity for Human Leukocyte Interferon with Monoclonal Antibodies" in Methods in Enzymology, Vol. 79 (Pestka, ed.), Academic Press, New York, 589-595.


2. Kelder et al. (1986) "A Sandwich Radioimmunoassay for Human IFNa" in Methods in Enzymology, Vol. 119 (Pestka, ed.), Academic Press, New York, 582-587.


3. Human IFN-α international reference standard provided by the NIH, reference no. Gxa01-901-535. Pestka (1986) "Interferon Standards and General Abbreviations" in Methods in Enzymology, Vol. 119 (Pestka, ed.), Academic Press, New York, 14-23.


4. Rubinstein et al. (1981) "Human Leukocyte Interferon: Isolation and Characterization of Several Molecular Forms," Arch. Biochem. Biophys. 210, 307-318.


5. Hobbs et al. (1982) "Purification and Characterization of Interferons from a Continuous Myeloblastic Cell Line," J. Biol. Chem. 257, 4071-4076.