Interferon Gamma Mouse mAb: Applications & Technical Guide

Interferon gamma Mouse mAb is a monoclonal antibody raised in mouse specifically targeting IFN-γ, offering high specificity for the detection and functional study of this critical pro-inflammatory cytokine across multiple immunological assay platforms.

Product SKU

Ab349543

Antibody Type

Mouse Monoclonal Antibody (mAb)

Target Antigen

Interferon Gamma (IFN-γ)

Host Species

Mouse

Primary Applications

ELISA, Western Blot, Flow Cytometry, Immunohistochemistry

Reference Clone/Catalog Note

Absea #KC-201 referenced

Background: Interferon Gamma and Its Biological Significance

Interferon gamma (IFN-γ) is a pleiotropic cytokine predominantly secreted by activated T helper 1 (Th1) cells, cytotoxic T lymphocytes (CTLs), and natural killer (NK) cells. It plays a central role in innate and adaptive immune responses, particularly in host defense against intracellular pathogens, tumor surveillance, and regulation of inflammatory processes.

IFN-γ signals through the JAK-STAT pathway by binding to its cognate receptor complex (IFNGR1/IFNGR2), leading to the phosphorylation of STAT1 and transcriptional activation of interferon-stimulated genes (ISGs). This results in upregulation of MHC class I and class II molecules, enhanced antigen presentation, and macrophage activation.

Given its pivotal role in immune regulation, accurate and reliable detection of IFN-γ is essential in research areas including autoimmune disease, infectious disease, cancer immunology, and vaccine development.

Product Description and Specificity of Ab349543

Ab349543 is a mouse-derived monoclonal antibody (mAb) directed against Interferon gamma (IFN-γ). Monoclonal antibodies offer the advantage of batch-to-batch consistency, defined epitope recognition, and high specificity compared to polyclonal alternatives, making them the preferred choice for quantitative and comparative immunological assays.

This antibody is referenced in association with Absea clone #KC-201, indicating a well-characterized clone with defined binding characteristics. Mouse monoclonal antibodies targeting IFN-γ are widely used in sandwich ELISA formats for quantitative cytokine measurement, as well as in intracellular cytokine staining (ICS) by flow cytometry to profile IFN-γ-producing cell populations.

Researchers should consult the product datasheet for confirmed species reactivity, recommended dilutions, and validated application details prior to experimental use.

Applications and Recommended Protocols

Enzyme-Linked Immunosorbent Assay (ELISA): Anti-IFN-γ monoclonal antibodies are routinely used as capture or detection antibodies in sandwich ELISA kits. For capture ELISA, coat high-binding microplates with the antibody at an appropriate concentration (typically 1–4 µg/mL in coating buffer, pH 9.6), block with 1–5% BSA or non-fat milk, then incubate with sample, followed by a biotinylated detection antibody and streptavidin-HRP conjugate.

Intracellular Cytokine Staining (ICS) / Flow Cytometry: Stimulate cells with appropriate activators (e.g., PMA/ionomycin or antigen-specific stimuli) in the presence of a protein transport inhibitor such as Brefeldin A or Monensin. After surface marker staining, fix and permeabilize cells, then incubate with the anti-IFN-γ mAb followed by a fluorochrome-conjugated secondary antibody if needed.

Western Blot: Separate cell lysates by SDS-PAGE, transfer to PVDF or nitrocellulose membrane, block, and probe with the anti-IFN-γ mAb at the recommended dilution. IFN-γ typically migrates at approximately 17 kDa (mature secreted form) under reducing conditions, though glycosylated forms may appear at higher apparent molecular weights.

Immunohistochemistry (IHC): Use on formalin-fixed paraffin-embedded (FFPE) or frozen tissue sections following standard antigen retrieval procedures. Optimize antibody concentration and incubation conditions empirically for each tissue type.

Storage, Handling, and Quality Considerations

Storage: Monoclonal antibodies should generally be stored at –20°C for long-term preservation or at 4°C for short-term use (typically up to one month). Avoid repeated freeze-thaw cycles, as these can denature the antibody and significantly reduce binding activity. Aliquot upon first thaw to minimize degradation.

Formulation: Antibodies are commonly supplied in PBS with a carrier protein such as BSA and a preservative such as sodium azide. Note that sodium azide is toxic and incompatible with peroxidase-based detection systems; dialysis or use of azide-free formulations is recommended for such applications.

Quality Control: Each lot of Ab349543 should be accompanied by lot-specific quality control data, including SDS-PAGE purity assessment and functional validation in at least one application. Researchers are advised to request the Certificate of Analysis (CoA) to confirm lot-specific performance specifications.

Working Concentrations: Optimal antibody concentrations vary by application and should be determined by titration. Consult the product datasheet for starting recommended concentrations and adjust based on signal-to-noise ratios observed in pilot experiments.

Frequently Asked Questions

See the FAQ section below for answers to common technical questions about working with Interferon Gamma Mouse mAb (Ab349543).

Frequently asked questions

What species reactivity does the Interferon Gamma Mouse mAb (Ab349543) have?

The species reactivity of Ab349543 should be confirmed by consulting the product datasheet or Certificate of Analysis, as this information is lot- and clone-specific. Commonly, anti-IFN-γ antibodies derived from mouse hosts may react with human, mouse, or other mammalian IFN-γ depending on the immunogen used. Contact Aladdin Scientific technical support for confirmed reactivity data.

Can Ab349543 be used for intracellular cytokine staining by flow cytometry?

Anti-IFN-γ monoclonal antibodies are commonly employed in intracellular cytokine staining (ICS) protocols. For ICS, cells must be stimulated, fixed, and permeabilized before antibody incubation to allow access to intracellular IFN-γ. Please verify that Ab349543 has been validated for ICS by reviewing the product datasheet or contacting Aladdin Scientific technical support, as not all mAb clones are suitable for all applications.

How should I handle and store Ab349543 to maintain optimal activity?

Store Ab349543 at –20°C for long-term use or at 4°C for up to one month. Avoid repeated freeze-thaw cycles by aliquoting the antibody upon first thaw. Keep the antibody away from light and ensure the vial is tightly sealed to prevent evaporation and contamination. Always consult the product datasheet for formulation-specific storage recommendations.

What is the expected molecular weight of IFN-γ detected by Western blot?

The mature secreted form of IFN-γ has an approximate molecular weight of 17 kDa under reducing SDS-PAGE conditions. However, due to N-linked glycosylation, the apparent molecular weight of the naturally occurring protein may range higher. Glycosylation patterns can vary depending on the cell type and species of origin.

References

1. Schroder K, Hertzog PJ, Ravasi T, Hume DA. Interferon-gamma: an overview of signals, mechanisms and functions. J Leukoc Biol. 2004;75(2):163-189. doi:10.1189/jlb.0603252
2. Platanias LC. Mechanisms of type-I- and type-II-interferon-mediated signalling. Nat Rev Immunol. 2005;5(5):375-386. doi:10.1038/nri1604
3. Absea Biotechnology. Product Reference: KC-201, Anti-Interferon gamma Monoclonal Antibody. Absea product documentation.

Products are supplied for research and development use only. Not for use in humans, animals, diagnosis, or therapy.