Revision 4

#10419Store at -20C

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, IHC-Bond, IHC-P, IF-IC, FC-FP, FC-L

REACTIVITY:

H

SENSITIVITY:

Endogenous

MW (kDa):

25

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q9Y5U5

Entrez-Gene Id:

8784

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50
IHC Leica Bond 1:100
Immunohistochemistry (Paraffin) 1:100
Immunofluorescence (Immunocytochemistry) 1:400
Flow Cytometry (Fixed/Permeabilized) 1:200
Flow Cytometry (Live) 1:200

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

For a carrier free (BSA and azide free) version of this product see product #24200.

Specificity / Sensitivity

GITR (D5V7P) Rabbit mAb recognizes endogenous levels of total GITR protein.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with recombinant protein specific to the amino terminus of human GITR protein.

Background

TNFRSF18, also known as glucocorticoid-induced tumor necrosis factor-receptor (TNFR)-related protein (GITR) and activation-inducible TNFR family receptor, encodes a type 1 membrane protein of the TNF-receptor superfamily (1). Three alternatively spliced transcript variants encoding distinct isoforms have been reported (2). GITR is an immune cell co-stimulatory receptor expressed constitutively at high levels on CD4+CD25+ T regulatory cells (Tregs), at low levels on naïve and memory T cells, and is induced upon T cell activation (3-5). Studies show GITR can also be induced on NK cells, macrophages, and DCs (3,4,6). Although GITR does not have intrinsic enzymatic activity, TNFSF18 (also known as GITRL) expressed on antigen presenting cells binds to GITR, resulting in recruitment of TNFR-associated factor family members and activation of the NF-κB pathway in T cells (7). GITR ligation has been shown to play a role in CD8+ T cell activation, cytotoxicity, and memory T cell survival (8-10). In the thymus, GITR is thought to play a key role in dominant immunological self-tolerance through thymic Treg differentiation and expansion (11). Of note, GITR ligation inhibits Treg suppressive function (12-13) and promotes effector T cell resistance to Treg suppression (14-15). Due to the combined effects on both Treg suppression and effector cell activation, GITR represents a unique opportunity for immunotherapeutic intervention in cancer (16).

  1. Nocentini, G. et al. (1997) Proc Natl Acad Sci U S A 94, 6216-21.
  2. Nocentini, G. et al. (2000) Cell Death Differ 7, 408-10.
  3. Shimizu, J. et al. (2002) Nat Immunol 3, 135-42.
  4. Nocentini, G. and Riccardi, C. (2009) Adv Exp Med Biol 647, 156-73.
  5. McHugh, R.S. et al. (2002) Immunity 16, 311-23.
  6. Hanabuchi, S. et al. (2006) Blood 107, 3617-23.
  7. Snell, L.M. et al. (2011) Immunol Rev 244, 197-217.
  8. Ronchetti, S. et al. (2007) J Immunol 179, 5916-26.
  9. Kim, I.K. et al. (2015) Nat Med 21, 1010-7.
  10. Snell, L.M. et al. (2012) J Immunol 188, 5915-23.
  11. Petrillo, M.G. et al. (2015) Autoimmun Rev 14, 117-26.
  12. Kanamaru, F. et al. (2004) J Immunol 172, 7306-14.
  13. Valzasina, B. et al. (2005) Blood 105, 2845-51.
  14. Stephens, G.L. et al. (2004) J Immunol 173, 5008-20.
  15. Nishikawa, H. et al. (2008) Cancer Res 68, 5948-54.
  16. Knee, D.A. et al. (2016) Eur J Cancer 67, 1-10.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation IHC-Bond: IHC Leica Bond IHC-P: Immunohistochemistry (Paraffin) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) FC-L: Flow Cytometry (Live)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

使用に関する制限

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Revision 4
#10419

GITR (D5V7P) Rabbit mAb

Western Blotting Image 1: GITR (D5V7P) Rabbit mAb Expand Image
Western blot analysis of extracts from HuT 102, human CD8+ T cells, and Jurkat cells using GITR (D5V7P) Rabbit mAb(upper), and β-Actin (D6A8) Rabbit mAb #8457 (lower). CD8+ T cells were purified from human blood and stimulated for 9 days using beads coated with CD3 and CD28 antibodies in the presence of human interleukin-2 (hIL-2) #8907 (6.7 ng/ml).
Immunoprecipitation Image 1: GITR (D5V7P) Rabbit mAb Expand Image
Immunoprecipitation of GITR protein from HuT 102 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is GITR (D5V7P) Rabbit mAb. Western blot analysis was performed using GITR (D5V7P) Rabbit mAb.
Immunohistochemistry Image 1: GITR (D5V7P) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human colon carcinoma using GITR (D5V7P) Rabbit mAb performed on the Leica® BOND Rx.
Immunohistochemistry Image 1: GITR (D5V7P) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded HuT 102 cell pellet (left, positive) or Jurkat cell pellet (right, negative) using GITR (D5V7P) Rabbit mAb.
Immunohistochemistry Image 2: GITR (D5V7P) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human colon carcinoma using GITR (D5V7P) Rabbit mAb.
Immunohistochemistry Image 3: GITR (D5V7P) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human lung adenocarcinoma using GITR (D5V7P) Rabbit mAb.
Immunohistochemistry Image 4: GITR (D5V7P) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human B-cell non-Hodgkin's lymphoma using GITR (D5V7P) Rabbit mAb.
Immunohistochemistry Image 5: GITR (D5V7P) Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human ovarian serous carcinoma using GITR (D5V7P) Rabbit mAb.
Immunofluorescence Image 1: GITR (D5V7P) Rabbit mAb Expand Image
Confocal immunofluorescent analysis of HuT 102 (left, positive) and Jurkat (right, negative) cells using GITR (D5V7P) Rabbit mAb (green). Blue = Hoechst 33342 #4082 (fluorescent DNA dye).
Flow Cytometry Image 1: GITR (D5V7P) Rabbit mAb Expand Image
Flow cytometric analysis of human peripheral blood mononuclear cells, untreated (left column) or treated with anti-CD3 (10 μg/ml, 72h) and anti-CD28 (5 μg/ml, 72h; right column), using GITR (D5V7P) Rabbit mAb (upper row) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype control #3900 (bottom row), and co-stained with a CD3 antibody. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody. Analysis was performed on CD3+ cells.
Flow Cytometry Image 2: GITR (D5V7P) Rabbit mAb Expand Image
Flow cytometric analysis of Jurkat cells (blue) and Hut102 cells (green), using GITR (D5V7P) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.