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Phospho-AMPK alpha 2 (S345) Recombinant Rabbit Monoclonal Antibody [JJ08-19]

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Specification

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Catalog# ET1701-37

Phospho-AMPK alpha 2 (S345) Recombinant Rabbit Monoclonal Antibody [JJ08-19]

Application

  • WB

  • IF-Tissue

  • IHC-P

  • FC

Reactivity

  • Human

  • Mouse

  • Rat

Conjugation

  • unconjugated

This product has been cited in peer reviewed publications, see list HERE

Overview

Product Name

Phospho-AMPK alpha 2 (S345) Recombinant Rabbit Monoclonal Antibody [JJ08-19]

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic phospho-peptide corresponding to residues surrounding Ser345 of Human AMPK alpha 2 aa 338-352 / 552.

Species Reactivity

Human, Mouse, Rat

Validated Applications

WB, IF-Tissue, IHC-P, FC

Molecular Weight

Predicted band size: 62 kDa

Positive Control

HEK-293 cell lysate, Mouse brain tissue lysate, Mouse heart tissue lysate, Rat brain tissue lysate, Rat heart tissue lysate, HEK-293, human skeletal muscle tissue, rat skeletal muscle tissue, rat heart tissue.

Conjugation

unconjugated

Clone Number

JJ08-19

RRID

AB_3070205

Product Features

Form

Liquid

Concentration

Storage Instructions

Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:500-1:2,000

  • IF-Tissue

  • 1:50-1:200

  • IHC-P

  • 1:50-1:200

  • FC

  • 1:1,000

Target

Function

Five-prime-AMP-activated protein kinase, known as AMPK, is a heterotrimeric complex that comprises of a catalytic α subunit, and regulatory β and γ. AMPK protects cells from stresses that cause ATP depletion by switching off ATP-consuming biosynthetic pathways. AMPK is activated by high AMP and low ATP via a mechanism involving allosteric regulation, promotion of phosphorylation by an upstream protein kinase known as AMPK kinase (AMPKK), and inhibition of dephosphorylation. Activated AMPK can phosphorylate and regulate in vivo hydroxymethylglutaryl-CoA reductase and acetyl-CoA carboxylase, which are key regulatory enzymes of sterol synthesis and fatty acid synthesis, respectively. The human AMPKα1 gene maps to chromosome 5p12 and encodes a 548 amino acid protein.

Background References

1. Hill K et al. Noise-Induced Loss of Hair Cells and Cochlear Synaptopathy Are Mediated by the Activation of AMPK. J Neurosci 36:7497-510 (2016).

2. Jiang S et al. AMPK-dependent regulation of GLP1 expression in L-like cells. J Mol Endocrinol 57:151-60 (2016).

Sequence Similarity

Belongs to the protein kinase superfamily. CAMK Ser/Thr protein kinase family. SNF1 subfamily.

Post-translational Modification

Ubiquitinated.; Phosphorylated at Thr-172 by STK11/LKB1 in complex with STE20-related adapter-alpha (STRADA) pseudo kinase and CAB39. Also phosphorylated at Thr-172 by CAMKK2; triggered by a rise in intracellular calcium ions, without detectable changes in the AMP/ATP ratio. CAMKK1 can also phosphorylate Thr-172, but at much lower level. Dephosphorylated by protein phosphatase 2A and 2C (PP2A and PP2C). Phosphorylated by ULK1; leading to negatively regulate AMPK activity and suggesting the existence of a regulatory feedback loop between ULK1 and AMPK. Dephosphorylated by PPM1A and PPM1B at Thr-172 (mediated by STK11/LKB1).

Subcellular Location

Nucleus, Cytoplasm.

UNIPROT

SwissProt: P54646 Human

SwissProt: Q8BRK8 Mouse

SwissProt: Q09137 Rat

Synonyms

5'-AMP-activated protein kinase catalytic subunit alpha-2 antibody

AAPK2_HUMAN antibody

ACACA kinase antibody

Acetyl-CoA carboxylase kinase antibody

AMPK alpha 2 chain antibody

AMPK subunit alpha-2 antibody

AMPK2 antibody

AMPKa2 antibody

AMPKalpha2 antibody

HMGCR kinase antibody

Expand

Images

  • Western blot analysis of Phospho-AMPK alpha 2 (S345) on different lysates with Rabbit anti-Phospho-AMPK alpha 2 (S345) antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>) at 1/1,000 dilution.<br /><br />Lane 1: HEK-293 cell lysate (20 µg/Lane)<br />Lane 2: Mouse brain tissue lysate (40 µg/Lane)<br />Lane 3: Mouse heart tissue lysate (40 µg/Lane)<br />Lane 4: Rat brain tissue lysate (40 µg/Lane)<br />Lane 5: Rat heart tissue lysate (40 µg/Lane)<br /><br />Predicted band size: 62 kDa<br />Observed band size: 62 kDa<br /><br />Exposure time: 3 minutes; ECL: K1801;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of Phospho-AMPK alpha 2 (S345) on different lysates with Rabbit anti-Phospho-AMPK alpha 2 (S345) antibody (ET1701-37) at 1/1,000 dilution.

    Lane 1: HEK-293 cell lysate (20 µg/Lane)
    Lane 2: Mouse brain tissue lysate (40 µg/Lane)
    Lane 3: Mouse heart tissue lysate (40 µg/Lane)
    Lane 4: Rat brain tissue lysate (40 µg/Lane)
    Lane 5: Rat heart tissue lysate (40 µg/Lane)

    Predicted band size: 62 kDa
    Observed band size: 62 kDa

    Exposure time: 3 minutes; ECL: K1801;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1701-37) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Flow cytometric analysis of HEK-293 cells labeling Phospho-AMPK alpha 2 (S345).<br /><br />Cells were fixed and permeabilized. Then stained with the primary antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor&trade; 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (<a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

    Flow cytometric analysis of HEK-293 cells labeling Phospho-AMPK alpha 2 (S345).

    Cells were fixed and permeabilized. Then stained with the primary antibody (ET1701-37, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

  • Immunohistochemical analysis of paraffin-embedded human skeletal muscle tissue with Rabbit anti-Phospho-AMPK alpha 2 (S345) antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>) at 1/50 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>) at 1/50 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human skeletal muscle tissue with Rabbit anti-Phospho-AMPK alpha 2 (S345) antibody (ET1701-37) at 1/50 dilution.

    The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1701-37) at 1/50 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded rat skeletal muscle tissue with Rabbit anti-Phospho-AMPK alpha 2 (S345) antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>) at 1/50 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>) at 1/50 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded rat skeletal muscle tissue with Rabbit anti-Phospho-AMPK alpha 2 (S345) antibody (ET1701-37) at 1/50 dilution.

    The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1701-37) at 1/50 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded rat heart tissue with Rabbit anti-Phospho-AMPK alpha 2 (S345) antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>) at 1/200 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1701-37" style="font-weight: bold;text-decoration: underline;">ET1701-37</a>) at 1/50 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded rat heart tissue with Rabbit anti-Phospho-AMPK alpha 2 (S345) antibody (ET1701-37) at 1/200 dilution.

    The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1701-37) at 1/50 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

Citation

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