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#35277 AMPK Substrate Antibody Sampler Kit

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2019年3月20日11時40分 現在
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#35277T1 Kit107,000
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AMPK substrate 製品一覧

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キット内容 容量 用途 種交差性 ホモロジー† 検出分子量 アイソタイプ
Phospho-AMPKα (Thr172) (D4D6D) Rabbit mAb #50081 20 µl WB, IP, IHC-P H, M, R C, Z, B, Pg 62 Rabbit IgG
AMPKα (D5A2) Rabbit mAb #5831 20 µl WB, IP H, M, R, Mk, B 62 Rabbit
Phospho-ULK1 (Ser555) (D1H4) Rabbit mAb #5869 20 µl WB, IP H, M R 140-150 Rabbit IgG
ULK1 (D8H5) Rabbit mAb #8054 20 µl WB, IP H, M, R, Mk 150 Rabbit IgG
Phospho-Raptor (Ser792) Antibody #2083 20 µl WB H, M, R 150 Rabbit
Raptor (24C12) Rabbit mAb #2280 20 µl WB, IP H, M, R, Mk 150 Rabbit
Beclin-1 (D40C5) Rabbit mAb #3495 20 µl WB, IP H, M, R, Mk 60 Rabbit IgG
Phospho-Beclin-1 (Ser93) (D9A5G) Rabbit mAb #14717 20 µl WB, IP H M, R, B, Dg, Pg 60 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody #7074 100 µl WB Goat

†Species predicted to react based on 100% sequence homology.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, IHC-P=Immunohistochemistry (Paraffin)
Reactivity Key: H=Human, M=Mouse, R=Rat, Mk=Monkey, B=Bovine

貯法
-20℃
社内データ

Western Blotting

Western Blotting

After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines, using Raptor (24C12) Rabbit mAb.

Western Blotting

Western Blotting

Western blot analysis of C2C12 or 293 cells, untreated or treated with AICAR (0.5 mM for 30 minutes) or oligomycin (0.5 μM for 30 minutes), using Phospho-Raptor (Ser792) Antibody (upper and lower left ) or Raptor Antibody #2280 (upper and lower right).

*Cross-reacting bands at 200 kDa.


Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Beclin-1 (D40C5) Rabbit mAb.

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa, K-562, C6, and Neuro-2a cells using AMPKα (D5A2) Rabbit mAb.

Western Blotting

Western Blotting

Western blot analysis of extracts from MCF7 cells, untreated or treated with oligomycin #9996 (0.5 μM, 30 minutes), and C2C12 cells, untreated or treated with hydrogen peroxide (10 mM, 5 minutes), using Phospho-ULK1 (Ser555) (D1H4) Rabbit mAb.


Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using ULK1 (D8H5) Rabbit mAb.

Western Blotting

Western Blotting

Western blot analysis of extracts from MCF7 and KARPAS-299 cells, untreated (-) or treated with carbonyl cyanide 3-chlorophenylhdrazone (CCCP, 100 μM, 2 hr; +), using Phospho-Beclin-1 (Ser93) (D9A5G) Rabbit mAb (upper) and Beclin-1 (D40C5) Rabbit mAb #3495 (lower). Cell line source: Dr Abraham Karpas at the University of Cambridge.

Western Blotting

Western Blotting

Western blot analysis of extracts from serum-starved NCI-H2228 cells, untreated (-) or treated with phenformin (5 mM, 1 hr; +), using Phospho-AMPKα (Thr172) (D4D6D) Rabbit mAb (upper) or AMPKα (D63G4) Rabbit mAb #5832 (lower).


Western Blotting

Western Blotting

Western blot analysis of wild-type (WT) and AMPKα1 and α2 knockout (KO) mouse embryonic fibroblasts (MEFs), untreated or treated with AICAR (2 mM for 1 hour), using Phospho-Raptor (Ser792) Antibody (upper) or Raptor Antibody #4978 (lower). (Image provided by Dr. Reuben Shaw, Salk Institute for Biological Studies).

*Cross-reacting bands at 60, 70 and 240 kDa

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence® Beclin-1 siRNA I #6222 (+) or SignalSilence® Beclin-1 siRNA II (+), using Beclin-1 (D40C5) XP® Rabbit mAb #3495 (upper) or α-Tubulin (11H10) Rabbit mAb #2125 (lower). The Beclin-1 (D40C5) XP® Rabbit mAb confirms silencing of Beclin-1 expression, while the α-Tubulin (11H10) Rabbit mAb is used to control for loading and specificity of Beclin-1 siRNA.

Western Blotting

Western Blotting

Western blot analysis of extracts from MCF7 cells, untreated or treated with oligomycin #9996 (0.5 µM, 30 minutes), using Phospho-ULK1 (Ser555) (D1H4) Rabbit mAb (left). Phospho-specificity is demonstrated by pre-incubating the antibody with phosphorylated (middle) or non-phosphorylated peptides (right) against a region surrounding Ser555 of ULK1.


Western Blotting

Western Blotting

Western blot analysis of extracts from wild-type MEF and ULK1 (-/-) MEF cells using ULK1 (D8H5) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower). MEF cells were kindly provided by Dr. Reuben Shaw (Salk Institute, La Jolla, CA).

Western Blotting

Western Blotting

Western blot analysis of extracts from 293T cells, untreated (-) or treated with carbonyl cyanide 3-chlorophenylhydrazone (CCCP, 100 μM, 2 hr; +) and mock transfected (-) or transfected with a construct expressing full-length human Beclin-1 (hBeclin-1; +), using Phospho-Beclin-1 (Ser93) (D9A5G) Rabbit mAb (upper) and Beclin-1 (D40C5) Rabbit mAb #3495 (lower).

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded NCI-H2228 cell pellets, untreated (left) or phenformin-treated (right), using Phospho-AMPKα (Thr172) (D4D6D) Rabbit mAb.


IP

IP

Immunoprecipitation of phospho-Beclin-1 (Ser93) from carbonyl cyanide 3-chlorophenyldydrazone (CCCP)-treated KARPAS-299 cells (100 μM, 2 hr) using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or Phospho-Beclin-1 (Ser93) (D9A5G) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot was performed using Phospho-Beclin-1 (Ser93) (D9A5G) Rabbit mAb. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb #3678 was used as a secondary antibody to avoid cross reactivity with IgG heavy chain. Cell line source: Dr Abraham Karpas at the University of Cambridge.

Western Blotting

Western Blotting

Western blot analysis of extracts from MCF7 cells, untreated (-) or treated with carbonyl cyanide 3-chlorophenylhydrazone (CCCP) (100 μM, 2 hr; +), using Phospho-ULK1 (Ser555) (D1H4) Rabbit mAb (upper) or ULK1 (D8H5) Rabbit mAb #8054 (lower).

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human colon carcinoma using Phospho-AMPKα (Thr172) (D4D6D) Rabbit mAb, control (left) or λ phosphatase-treated (right).


IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human ovarian carcinoma using Phospho-AMPKα (Thr172) (D4D6D) Rabbit mAb.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Phospho-AMPKα (Thr172) (D4D6D) Rabbit mAb in the presence of control peptide (left) or antigen-specific peptide (right).

バックグラウンド

AMP-activated protein kinase (AMPK) is highly conserved from yeast to plants and animals and plays a key role in the regulation of energy homeostasis (1). AMPK is a heterotrimeric complex composed of a catalytic α subunit and regulatory β and γ subunits, each of which is encoded by two or three distinct genes (α1, 2; β1, 2; γ1, 2, 3) (2). The kinase is activated by an elevated AMP/ATP ratio due to cellular and environmental stress, such as heat shock, hypoxia, and ischemia (1). The tumor suppressor LKB1, in association with accessory proteins STRAD and MO25, phosphorylates AMPKα at Thr172 in the activation loop, and this phosphorylation is required for AMPK activation (3-5).

AMPK phosphorylates a number of targets controlling cellular processes such as metabolism, cell growth, and autophagy (6). It suppresses the activity of the mammalian target of rapamycin (mTOR), that plays a key role in promoting cell growth. The regulatory associated protein of mTOR (Raptor) was identified as an mTOR binding partner that mediates mTOR signaling to downstream targets (7,8). Raptor binds to mTOR substrates, including 4E-BP1 and p70 S6 kinase, through their TOR signaling (TOS) motifs and is required for mTOR-mediated phosphorylation of these substrates (9,10). AMPK directly phosphorylates Raptor at Ser722/Ser792, and this phosphorylation is essential for inhibition of the raptor-containing mTOR complex 1 (mTORC1) and induces cell cycle arrest when cells are stressed for energy (11). AMPK also promotes autophagy by directly phosphorylating ULK1 (11,12). ULK1 is a Ser/Thr kinase required for the Initiation and formation of the autophagosome. AMPK, activated during low nutrient conditions, directly phosphorylates ULK1 at multiple sites including Ser317, Ser555, and Ser777 (11,12). Conversely, mTOR, which is a regulator of cell growth and an inhibitor of autophagy, phosphorylates ULK1 at Ser757 and disrupts the interaction between ULK1 and AMPK (11). AMPK can also directly phosphorylate Beclin-1, a component of the complex downstream of ULK1 in autophagosome formation that activates the class III phosphatidylinositol 3-kinase VPS34. AMPK phosphorylates Beclin-1 at Ser93 and Ser96 residues in human, which correspond to murine Ser91 and Ser94 (14).

使用例
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関連製品
14717   Phospho-Beclin-1 (Ser93) (D9A5G) Rabbit mAb
2083   Phospho-Raptor (Ser792) Antibody
2280   Raptor (24C12) Rabbit mAb
3495   Beclin-1 (D40C5) Rabbit mAb
50081   Phospho-AMPKα (Thr172) (D4D6D) Rabbit mAb
5831   AMPKα (D5A2) Rabbit mAb
5869   Phospho-ULK1 (Ser555) (D1H4) Rabbit mAb
7003   20X LumiGLO® Reagent and 20X Peroxide
7071   Phototope-HRP Western Blot Detection System, Anti-rabbit IgG, HRP-linked Antibody
7074   Anti-rabbit IgG, HRP-linked Antibody
7727   Biotinylated Protein Ladder Detection Pack
8054   ULK1 (D8H5) Rabbit mAb

Tween is a registered trademark of ICI Americas, Inc.
SignalStain is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
LumiGLO is a registered trademark of Kirkegaard & Perry Laboratories.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.

本製品は試験研究用です。

AMPK Substrate Antibody Sampler Kit

Immune Cell Signaling Pathways

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