Revision 7
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, IHC-P, IF-F, IF-IC, FC-FP

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

46, 48

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q92597

Entrez-Gene Id:

10397

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunohistochemistry (Paraffin) 1:250 - 1:1000
Immunofluorescence (Frozen) 1:50 - 1:100
Immunofluorescence (Immunocytochemistry) 1:200 - 1:800
Flow Cytometry (Fixed/Permeabilized) 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 #89166.

Specificity / Sensitivity

Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb detects endogenous levels of NDRG1 when phosphorylated at Thr346. This antibody likely cross-reacts with other conserved phosporylation sites on NDRG1 at positions Thr356 and Thr366.

Species Reactivity:

Human, Mouse, Rat, Monkey

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Thr346 of mouse NDRG1 protein.

Background

N-myc downstream-regulated gene 1 (NDRG1), also termed Cap43, Drg1, RTP/rit42, and Proxy-1, is a member of the NDRG family, which is composed of four members (NDRG1-4) that function in growth, differentiation, and cell survival (1-5). NDRG1 is ubiquitously expressed and highly responsive to a variety of stress signals, including DNA damage (4), hypoxia (5), and elevated levels of nickel and calcium (2). Expression of NDRG1 is elevated in N-myc defective mice and is negatively regulated by N- and c-myc (1,6). During DNA damage, NDRG1 is induced in a p53-dependent fashion and is necessary for p53-mediated apoptosis (4,7). Research studies have shown that NDRG1 may also play a role in cancer progression by promoting differentiation, inhibiting growth, and modulating metastasis and angiogenesis (3,4,6,8,9). Nonsense mutation of the NDRG1 gene has been shown to cause hereditary motor and sensory neuropathy-Lom (HMSNL), which is supported by studies demonstrating the role of NDRG1 in maintaining myelin sheaths and axonal survival (10,11). NDRG1 is upregulated during mast cell maturation and its deletion leads to attenuated allergic responses (12). Both NDRG1 and NDRG2 are substrates of SGK1, although the precise physiological role of SGK1-mediated phosphorylation is not known (13). NDRG1 is phosphorylated by SGK1 at Thr328, Ser330, Thr346, Thr356, and Thr366. Phosphorylation by SGK1 primes NDRG1 for phosphorylation by GSK-3.

Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb is directed at a site that was identified at Cell Signaling Technology (CST) using PhosphoScan®, CST's LC-MS/MS platform for modification site discovery. Phosphorylation at Thr346 was discovered using an Akt substrate antibody and was shown to be induced by insulin treatment in multiple cell lines. Please visit PhosphoSitePlus®, CST's modification site knowledgebase, at www.phosphosite.org for more information.

  1. Shimono, A. et al. (1999) Mech Dev 83, 39-52.
  2. Zhou, D. et al. (1998) Cancer Res 58, 2182-9.
  3. van Belzen, N. et al. (1997) Lab Invest 77, 85-92.
  4. Kurdistani, S.K. et al. (1998) Cancer Res 58, 4439-44.
  5. Park, H. et al. (2000) Biochem Biophys Res Commun 276, 321-8.
  6. Li, J. and Kretzner, L. (2003) Mol Cell Biochem 250, 91-105.
  7. Stein, S. et al. (2004) J Biol Chem 279, 48930-40.
  8. Maruyama, Y. et al. (2006) Cancer Res 66, 6233-42.
  9. Nishio, S. et al. (2008) Cancer Lett 264, 36-43.
  10. Kalaydjieva, L. et al. (2000) Am J Hum Genet 67, 47-58.
  11. Okuda, T. et al. (2004) Mol Cell Biol 24, 3949-56.
  12. Taketomi, Y. et al. (2007) J Immunol 178, 7042-53.
  13. Murray, J.T. et al. (2004) Biochem J 384, 477-88.

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 nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IHC-P: Immunohistochemistry (Paraffin) IF-F: Immunofluorescence (Frozen) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized)

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.
XP is a registered trademark of Cell Signaling Technology, Inc.
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

法的な権限を与えられたCSTの担当者が署名した書面によって別途明示的に合意された場合を除き、 CST、その関連会社または代理店が提供する製品には以下の条件が適用されます。お客様が定める条件でここに定められた条件に含まれるものを超えるもの、 または、ここに定められた条件と異なるものは、法的な権限を与えられたCSTの担当者が別途書面にて受諾した場合を除き、拒絶され、 いかなる効力も効果も有しません。

研究専用 (For Research Use Only) またはこれに類似する表示がされた製品は、 いかなる目的についても FDA または外国もしくは国内のその他の規制機関により承認、認可または許可を受けていません。 お客様は製品を診断もしくは治療目的で使用してはならず、また、製品に表示された内容に違反する方法で使用してはなりません。 CST が販売または使用許諾する製品は、エンドユーザーであるお客様に対し、使途を研究および開発のみに限定して提供されるものです。 診断、予防もしくは治療目的で製品を使用することまたは製品を再販売 (単独であるか他の製品等の一部であるかを問いません) もしくはその他の商業的利用の目的で購入することについては、CST から別途許諾を得る必要があります。 お客様は以下の事項を遵守しなければなりません。(a) CST の製品 (単独であるか他の資材と一緒であるかを問いません) を販売、使用許諾、貸与、寄付もしくはその他の態様で第三者に譲渡したり使用させたりしてはなりません。また、商用の製品を製造するために CST の製品を使用してはなりません。(b) 複製、改変、リバースエンジニアリング、逆コンパイル、 分解または他の方法により製品の構造または技術を解明しようとしてはなりません。また、 CST の製品またはサービスと競合する製品またはサービスを開発する目的で CST の製品を使用してはなりません。(c) CST の製品の商標、商号、ロゴ、特許または著作権に関する通知または表示を除去したり改変したりしてはなりません。(d) CST の製品をCST 製品販売条件(CST’s Product Terms of Sale) および該当する書面のみに従って使用しなければなりません。(e) CST の製品に関連してお客様が使用する第三者の製品またはサービスに関する使用許諾条件、 サービス提供条件またはこれに類する合意事項を遵守しなければなりません。

Revision 7
#5482

Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb

Western Blotting Image 1: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from C2C12 cells, untreated or treated with LY294002 #9901, insulin, or both using Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb (upper) or total NDRG1 Antibody #5196 (lower).
Western Blotting Image 2: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from A431 cells treated with hEGF #8916 (100 ng/ml, 30 minutes), with or without calf intestinal phosphatase and λ-phosphatase, using Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb (upper) or total NDRG1 Antibody #5196 (lower).
Western Blotting Image 3: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Western blot analysis of HeLa cells treated with hEGF #8916 (100 ng/ml, 30 minutes), mock transfected or transfected with SignalSilence® NDRG1 siRNA I #6245 or SignalSilence® NDRG1 siRNA II #6257, using Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb (upper) or β-Tubulin (9F3) Rabbit mAb #2128 (lower).
Immunohistochemistry Image 1: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Immunohistochemical analysis on SignalSlide® Phospho-Akt (Ser473) IHC Controls #8101 [paraffin-embedded LNCaP cell pellets, control (left) or LY294002-treated (right)] using NDRG1 Antibody #5196 (top) or Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb (bottom).
Immunohistochemistry Image 2: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-emebedded mouse colon using Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb.
Immunohistochemistry Image 3: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-embedded human cervical carcinoma, control (left) or λ phosphatase-treated (right), using Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb.
Immunohistochemistry Image 4: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Immunohistochemical analysis of paraffin-emebedded human lung carcinoma using Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb.
Immunofluorescence Image 1: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of fixed frozen mouse colon labeled with Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb #5482 (green). Free secondary binding sites were then blocked with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 prior to colabeling with α-Smooth Muscle Actin (D4K9N) XP® Rabbit mAb (Alexa Fluor® 555 Conjugate) #60839 (yellow), DyLight 650 Phalloidin #12956 (red), and DAPI #4083 (blue).
Immunofluorescence Image 2: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of fixed frozen mouse kidney labeled with Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb #5482 (left, green) and colabeled DyLight 650 Phalloidin #12956 (right, red) and DAPI #4083 (right, blue).
Immunofluorescence Image 3: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of fixed frozen mouse striatum labeled with Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb #5482 (green). Free secondary binding sites were then blocked with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 prior to colabeling with Myelin Basic Protein (D8X4Q) XP® Rabbit mAb (Alexa Fluor® 555 Conjugate) #84987 (yellow), GFAP (GA5) Mouse mAb (Alexa Fluor® 647 Conjugate) #3657 (red), and DAPI #4083 (blue).
Immunofluorescence Image 1: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of C2C12 cells, treated with LY294002 #9901 (left) or insulin (right), using Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Flow Cytometry Image 1: Phospho-NDRG1 (Thr346) (D98G11) XP® Rabbit mAb Expand Image

Flow cytometric analysis of Jurkat cells, untreated (green) or treated for 2 hr with LY294002 #9901 (50 µM), Wortmannin #9951 (1 µM), and U0126 #9903 (10 μM) (blue) using Phospho-NDRG1 (Thr346) (D98G11) XP® 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.