Cell Signaling Technology
XP Monoclonal Antibody

Product Pathways - Growth Factors/Cytokines

IFN-γ (D3H2) XP® Rabbit mAb #8455

Applications Reactivity Sensitivity MW (kDa) Isotype
W IP IF-IC F H Endogenous 17, 19, 23 Rabbit IgG

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry
Reactivity Key:  H=Human
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

* Product-specific protocol.

Specificity / Sensitivity

IFN-γ (D3H2) XP® Rabbit mAb recognizes endogenous levels of total IFN-γ protein.

Source / Purification

Monoclonal antibody is produced by immunizing animals with recombinant human IFN-γ protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from NK-92 cells, untreated (-) or treated (+) with TPA #4174 (80 nM, 5 hr), Ionomycin #9995 (3 μM, 5 hr), and Brefeldin A #9972 (300 ng/mL, last 4 hr of stimulation), using IFN-γ (D3H2) XP® Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).

IP

IP

Immunoprecipitation of IFN-γ from NK-92 cell extracts, treated with TPA #4174 (80 nM, 5 hr), Ionomycin #9995 (3 μM, 5 hr), and Brefeldin A #9972 (300 ng/mL, last 4 hr of stimulation), using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or IFN-γ (D3H2) XP® Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using IFN-γ (D3H2) XP® Rabbit mAb.

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of human peripheral blood cells, untreated (left) or treated (right) with TPA #4174 (40 nM, 4 hr), Ionomycin #9995 (2 μM, 4 hr), and Brefeldin A #9972 (1 μg/mL, last 3 hr of stimulation), using a CD3 antibody and IFN-γ (D3H2) XP® Rabbit mAb. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) #4414 was used as a secondary antibody. Analysis was performed on cells in the lymphocyte gate.


IF-IC

IF-IC

Confocal immunofluorescent analysis of NK-92 cells, treated with TPA #4174 and Ionomycin #9995 (80 nM and 3 μM, 4 hr; left) or untreated (right), using IFN-γ (D3H2) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

Background

IFN-γ plays key roles in both the innate and adaptive immune response. IFN-γ activates the cytotoxic activity of innate immune cells, such as macrophages and NK cells (1,2). IFN-γ production by NK cells and antigen presenting cells (APCs) promotes the cell mediated adaptive immunity by inducing IFN-γ production by T lymphocytes, increasing class I and class II MHC expression, and enhancing peptide antigen presentation (1). The anti-viral activity of IFN-γ is due to its induction of PKR and other regulatory proteins. Binding of IFN-γ to the IFNGR1/IFNGR2 complex promotes dimerization of the receptor complexes to form the (IFNGR1/IFNGR2)2 -IFN-γ dimer. Binding induces a conformational change in receptor intracellular domains and signaling involves Jak1, Jak2, and Stat1 (3). The critical role of IFN-γ in amplification of immune surveillance and function is supported by increased susceptibility to pathogen infection by IFN-γ or IFNGR knockout mice and in humans with inactivating mutations in IFNGR1 or IFNGR2. IFN-γ also appears to have a role in atherosclerosis (4).

  1. Schroder, K. et al. (2004) J Leukoc Biol 75, 163-89.
  2. Martinez, F.O. et al. (2009) Annu Rev Immunol 27, 451-83.
  3. Kotenko, S.V. et al. (1995) J Biol Chem 270, 20915-21.
  4. McLaren, J.E. and Ramji, D.P. (2009) Cytokine Growth Factor Rev 20, 125-35.

Application References

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Companion Products

For Research Use Only. Not For Use In Diagnostic Procedures.

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