Human IFN-alpha All Subtype Quantikine ELISA Kit

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Human IFN-Alpha All Subtype Quantikine Spiked Linearity Competitor Comparison
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Human IFN-alpha All Subtype Quantikine ELISA Kit Summary

Assay Length
4.5 hours
Sample Type & Volume Required Per Well
Cell Culture Supernates (50 uL), Serum (50 uL), EDTA Plasma (50 uL), Heparin Plasma (50 uL)
Sensitivity
4.8 pg/mL
Assay Range
15.6 - 1,000 pg/mL (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma)
Specificity
Natual and recombinant human IFN- alpha.
Cross-reactivity
< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.
Interference
No significant interference observed with available related molecules.

Product Summary

The Quantikine® Human IFN-alpha All Subtype Immunoassay is a 4.5 hour solid phase ELISA designed to measure all human IFN-alpha subtypes in cell culture supernates, serum, and plasma. It contains HEK293-expressed recombinant human IFN-alpha 2a and has been shown to accurately quantitate the recombinant factor. Results obtained using natural human IFN-alpha 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 human IFN-alpha subtypes.

Precision

Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested twenty times on one plate to assess intra-assay precision
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in twenty separate assays to assess inter-assay precision. Assays were performed by at least three technicians using two lots of components

Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma

Intra-Assay Precision Inter-Assay Precision
Sample 1 2 3 1 2 3
n 20 20 20 20 20 20
Mean (pg/mL) 178 340 698 185 354 737
Standard Deviation 3.62 8.35 20.9 9.5 20.5 41
CV% 2 2.5 3 5.1 5.8 5.6

Recovery

The recovery of human IFN-alpha spiked to levels throughout the range of the assay in various matrices was evaluated.

Sample Type Average % Recovery Range %
Cell Culture Media (n=4) 101 96-103
EDTA Plasma (n=4) 92 89-96
Heparin Plasma (n=4) 91 88-96
Serum (n=4) 89 83-94

Linearity

To assess the linearity of the assay, samples containing and/or spiked with high concentrations of human IFN-alpha were serially diluted with calibrator diluent to produce samples with values within the dynamic range of the assay.
Human IFN-alpha  All Subtype ELISA Linearity

Scientific Data

Human IFN-Alpha All Subtype Quantikine Spiked Linearity Competitor Comparison IFN-alpha is spiked at high concentration in various matrices and diluted with appropriate Calibrator Diluent to produce samples with values within the dynamic range of the assay. R&D Systems linearity is between 105%-112% compared to 114%-134% and 109%-129% for the top competitors.

Human IFN-Alpha All Subtype Spiked Recovery Competitor Comparison IFN-alpha is spiked at three known concentrations throughout the range of the assay and run to measure response of the spiked sample matrix. R&D Systems serum recovery is 104% compared to 74% and 163% for the top competitors.

Human IFN-alpha All Subtype ELISA Standard Curve

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Preparation and Storage

Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: IFN-alpha

There are 3 major classes of interferons (IFNs): Type I, Type II and Type III. Interferon alpha (IFN-alpha ), along with IFN-beta, IFN-δ, IFN-epsilon, IFN-kappa, IFN-omega and IFN-tau are all Type I IFNs (1). The sole type II IFN is IFN-gamma. Type III IFNs include IFN-lambda 1, IFN-lambda 2, IFN-lambda 3 and IFN-lambda 4 (2). As a part of the innate immune response, Type I IFNs are rapidly induced in response to viral nucleic acids such as double stranded DNA or RNA (dsDNA, dsRNA) and single stranded RNA (ssRNA), viral glycoproteins, microbial cytosinephosphate-guanosine (CpG) DNA, DNA damage, and chromosomal instability (3,4). 

IFN-alpha subtypes are well described (5-9). There are 15 human IFN-alpha subtypes with 80% amino acid identity (10). The number of IFN-alpha subtypes varies by species with 6 equine subtypes, 17 porcine subtypes, 14 bovine subtypes, and 9 canine subtypes known currently. Human IFN-alpha subtypes include: IFN-alpha 1a, IFN-alpha 1b, IFN-alpha 2a, IFN-alpha 2b, IFN-alpha 4a, IFN-alpha 4b, IFN-alpha 5, IFN-alpha 6 IFN-alpha 7, IFN-alpha 8, IFN-alpha 10, IFN-alpha 14, IFN-alpha 16, IFN-alpha 17, and IFN-alpha 21. Although there is one known heterodimeric IFN-alpha receptor (IFN-alpha R, described below), each IFN-alpha subtype has been correlated with differing biological activities (7). Variability in biological responses can be attributed to differences in binding affinity and duration, receptor density, feedback responses and intracellular characteristics (11). IFN-alpha responses have been described as robust, especially in the context of viral infection responsiveness by all cells or tunable in a cell type specific manner. 
IFN-alpha signaling is well characterized (2, 5,11,12,13). IFN-alpha is a ligand for IFN alpha R, which includes two subunits IFN alpha R1 and IFN alpha R2. IFN-alpha ligand binding to the ubiquitously expressed IFN alpha R1 triggers a conformational change which allows for the heterodimerization of IFN alpha R1 and IFN alpha R2 (10). Heterodimerization results in the cross phosphorylation of the Janus-activated Kinase 1 (JAK1) on IFN alpha R2 and tyrosine kinase 2 (TYK2) on IFN alpha R1 respectively, as well as the intracellular domain of IFN alpha R1 and IFN alpha 2 (9). The transcription factors Signal transducer and activator of transcription (STAT) 1 and 2 are subsequently recruited to IFN alpha R via their Src homology 2 (SH2) domain and phosphorylated. In the canonical IFN-alpha signaling pathway, phosphoSTAT1/STAT2 heterodimers associate with Interferon Regulatory Factor 9 (IRF9) to form Interferon Stimulated Gene Factor 3 (ISGF3), which translocates into the nucleus to bind to gamma-activated sequences (GASs) or interferon- stimulated response elements (ISREs). This stimulates the transcription of interferonstimulated genes (ISGs). STAT 2 homodimers and monomers can also associate with IRF-9 to form ISGF3-like complex, which also binds to ISRE to stimulate the transcription of ISGs (9). 
Although IFN-alpha is most commonly associated with viral infections, it has been associated with other pathological events. Type 1 interferons represent a standard of care for suppressing Hepatitis B (HBV) or C (HCV) (14). However, it has been associated with neuropsychiatric symptoms such as depression, anhedonia, anxiety and cognitive impairment (15). The role of IFN-alpha in cancer is complex as well. For example, in the context of inflammatory breast cancer, IFN-alpha is upregulated. Although IFN-alpha has been correlated with cellular senescence and apoptosis, some subtypes have been correlated with increased cellular migration and drug resistance.

Long Name:
Interferon alpha
Entrez Gene IDs:
3440 (Human)
Alternate Names:
IFNA; IFNA2; IFNA2b; IFNalpha; IFN-alpha; IFN-alphaA; INFA2; interferon alpha 2; LeIF D

Citations for Human IFN-alpha All Subtype Quantikine ELISA Kit

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

6 Citations: Showing 1 - 6
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  1. Complete substitution with modified nucleotides suppresses the early interferon response and increases the potency of self-amplifying RNA
    Authors: McGee, JE;Kirsch, JR;Kenney, D;Chavez, E;Shih, TY;Douam, F;Wong, WW;Grinstaff, MW;
    bioRxiv : the preprint server for biology
    Species: Human
    Sample Types: Cell Culture Supernates
  2. The IL-4/IL-13 signaling axis promotes prostatic fibrosis
    Authors: Q D'Arcy, M Gharaee-Ke, A Zhilin-Rot, JA Macoska
    PLoS ONE, 2022-10-06;17(10):e0275064.
    Species: Human
    Sample Types: Cell Culture Supernates
  3. Mechanisms regulating transitory suppressive activity of neutrophils in newborns: PMNs-MDSCs in newborns
    Authors: M Perego, S Fu, Y Cao, A Kossenkov, M Yao, K Alicea-Tor, W Liu, Z Jiang, Z Chen, SY Fuchs, J Zhou, DI Gabrilovic
    Oncogene, 2022-06-21;0(0):.
    Species: Human
    Sample Types: Plasma
  4. Epigenetic Regulation of IFI44L Expression in Monocytes Affects the Functions of Monocyte-Derived Dendritic Cells in Systemic Lupus Erythematosus
    Authors: S Luo, R Wu, Q Li, G Zhang
    Journal of Immunology Research, 2022-05-10;2022(0):4053038.
    Species: Human
    Sample Types: Serum
  5. In Vitro SARS-CoV-2 Infection of Microvascular Endothelial Cells: Effect on Pro-Inflammatory Cytokine and Chemokine Release
    Authors: M Dolci, L Signorini, S D'Alessand, F Perego, S Parapini, M Sommariva, D Taramelli, P Ferrante, N Basilico, S Delbue
    International Journal of Molecular Sciences, 2022-04-06;23(7):.
    Species: Human
    Sample Types: Cell Culture Supernates
  6. Mitochondria-localised ZNFX1 functions as a dsRNA sensor to initiate antiviral responses through MAVS
    Authors: Y Wang, S Yuan, X Jia, Y Ge, T Ling, M Nie, X Lan, S Chen, A Xu
    Nat. Cell Biol., 2019-11-04;21(11):1346-1356.
    Species: Mouse
    Sample Types: Cell Culture Supernates

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Reviews for Human IFN-alpha All Subtype Quantikine ELISA Kit

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Human IFN-alpha All Subtype Quantikine ELISA Kit
By Anonymous on 12/04/2023
Sample Tested: B cell enriched peripheral blood mononuclear cells (PBMC)

Good dynamic range.
cell supernatent with good linearity (1:2 to 1:16)


Human IFN-alpha All Subtype Quantikine ELISA Kit
By Anonymous on 12/01/2023
Sample Tested: Cell Culture Supernates

I used 40 ul of the samples only.