Published: Vol 4, Iss 10, May 20, 2014 DOI: 10.21769/BioProtoc.1130 Views: 11402
Reviewed by: Anonymous reviewer(s)
Protocol Collections
Comprehensive collections of detailed, peer-reviewed protocols focusing on specific topics
Related protocols
Protein Pull-down Assay Using HiBiT-tag-dependent Luciferase Activity Measurement
Masashi Arakawa and Eiji Morita
Mar 20, 2023 1403 Views
Mass Spectrometry-based Lipidomics, Lipid Bioenergetics, and Web Tool for Lipid Profiling and Quantification in Human Cells
Liang Cui [...] Kuan Rong Chan
Aug 20, 2023 1585 Views
Cell-Sonar, an Easy and Low-cost Method to Track a Target Protein by Expression Changes of Specific Protein Markers
Sabrina Brockmöller [...] Simone Rothmiller
Feb 5, 2025 483 Views
Abstract
Type I interferons (IFN-α/β) play an important role in host resistance to viral infections. Signaling through the JAK-STAT pathway, IFN-α/β stimulates response elements (ISRE) in the promoters of ISG to regulate their expression (reviewed in Reference 2). This method was adapted from InvivoGen to specifically detect and quantify IFN-α/β secreted in response to virus infection. HEK-Blue™ IFN-α/β cells were generated by stably introducing the human STAT2 and IRF9 genes into HEK293 cells to obtain a fully active type I IFN signaling pathway. The activation of this pathway is made detectable by the addition of a reporter gene expressing a secreted embryonic alkaline phosphatase (SEAP) under the control of the ISG54 promoter. ISG54 is a well-known ISG activated through an ISRE-dependent mechanism by type I IFNs.
Materials and Reagents
Equipment
Procedure
IFN (1) | IFN (1) | IFN (1) | Sample 1 | Sample 1 | Sample 1 | | | | | | |
IFN (2) | IFN (2) | IFN (2) | Sample 2 | Sample 2 | Sample 2 | | | | | | |
IFN (3) | IFN (3) | IFN (3) | Sample 3 | Sample 3 | Sample 3 | | | | | | |
IFN (4) | IFN (4) | IFN (4) | Sample 4 | Sample 4 | Sample 4 | | | | | | |
IFN (5) | IFN (5) | IFN (5) | Sample 5 | Sample 5 | Sample 5 | | | | | | |
IFN (6) | IFN (6) | IFN (6) | Mock | Mock | Mock | | | | | | |
IFN (7) | IFN (7) | IFN (7) | Neg.c. 1 | Neg.c. 1 | Neg.c. 1 | | | | | | |
IFN (8) | IFN (8) | IFN (8) | Neg.c. 2 | Neg.c. 2 | Neg.c. 2 | | | | | | |
1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 |
10,000 U/ml | 3,333 U/ml | 1,111 U/ml | 370 U/ml | 123 U/ml | 41 U/ml | 13.7 U/ml | 4.6 U/ml |
75 µl of 104 IU/ml | 50 µl DMEM | 50 µl DMEM | 50 µl DMEM | 50 µl DMEM | 50 µl DMEM | 50 µl DMEM | 50 µl DMEM |
Notes
Acknowledgments
This protocol was published in the original paper by Lulla et al. (2013). This work was supported by Estonian Science Foundation grants 7501, 7407, and9421; target financing project SF0180087s08; and the European Union through the European Regional Development Fund via the Center of Excellence in Chemical Biology.
References
Article Information
Copyright
© 2014 The Authors; exclusive licensee Bio-protocol LLC.
How to cite
Lulla, V., Merits, A. and Lulla, A. (2014). IFN-α/β Detection Assay Using Sensor Cell Lines. Bio-protocol 4(10): e1130. DOI: 10.21769/BioProtoc.1130.
Category
Microbiology > Microbe-host interactions > Virus
Biochemistry > Protein > Expression
Do you have any questions about this protocol?
Post your question to gather feedback from the community. We will also invite the authors of this article to respond.
Tips for asking effective questions
+ Description
Write a detailed description. Include all information that will help others answer your question including experimental processes, conditions, and relevant images.
Share
Bluesky
X
Copy link