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Phospho-EEF2 (T56) Recombinant Rabbit Monoclonal Antibody [PSH06-36] - BSA and Azide free

Usd: 649

Specification

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Catalog# HA751075

Phospho-EEF2 (T56) Recombinant Rabbit Monoclonal Antibody [PSH06-36] - BSA and Azide free

Application

  • WB

  • IHC-P

  • IF-Cell

Reactivity

  • Human

  • Mouse

  • Rat

With BSA and Azide
Conjugation

  • unconjugated

Overview

Product Name

Phospho-EEF2 (T56) Recombinant Rabbit Monoclonal Antibody [PSH06-36] - BSA and Azide free

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic phosphopeptide corresponding to residues surrounding Thr56 of human EEF2.

Species Reactivity

Human, Mouse, Rat

Validated Applications

WB, IHC-P, IF-Cell

Molecular Weight

Predicted band size: 95 kDa

Positive Control

HeLa starved for 24 hours then add 10μM forskolin for 1 hour cell lysate, NIH/3T3 cell lysate, NIH/3T3 starved for 24 hours then add 10μM forskolin for 1 hour cell lysate, PC-12 treated with 20μM forskolin for 1 hour then add 100nM Calyculin A for 30 minutes cell lysate, HepG2 cell lysate, rat liver tissue, HeLa cells starved for 24 hours then add 10μM forskolin for 1 hour, PC-12 cells treated with 20μM forskolin for 1 hour then add 100nM Calyculin A for 30 minutes.

Conjugation

unconjugated

Clone Number

PSH06-36

Product Features

Form

Liquid

Concentration

1mg/ml

Storage Instructions

Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.

Storage Buffer

1*PBS (pH7.4).

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:2,000

  • IHC-P

  • 1:500

  • IF-Cell

  • 1:100

Target

Function

Eukaryotic elongation factor 2 is a protein that in humans is encoded by the EEF2 gene. It is the archaeal and eukaryotic counterpart of bacterial EF-G. This gene encodes a member of the GTP-binding translation elongation factor family. This protein is an essential factor for protein synthesis. It promotes the GTP-dependent translocation of the ribosome. This protein is completely inactivated by EF-2 kinase phosphorylation. aEF2/eEF2 found in most archaea and eukaryotes, including humans, contains a post translationally modified histidine diphthamide. It is the target of diphtheria toxin (from Corynebacterium diphtheriae), and exotoxin A (from Pseudomonas aeruginosa). The inactivation of EF-2 by toxins inhibits protein production in the host, causing symptoms due to loss of function in affected cells.

Background References

1. Chen Q et al. Inhibition of LDHA to induce eEF2 release enhances thrombocytopoiesis. Blood. 2022 May

2. Li W et al. Fluoxetine regulates eEF2 activity (phosphorylation) via HDAC1 inhibitory mechanism in an LPS-induced mouse model of depression. J Neuroinflammation. 2021 Feb

Subcellular Location

Cytoplasm, Nucleus.

UNIPROT

SwissProt: P13639 Human

SwissProt: P58252 Mouse

SwissProt: P05197 Rat

Synonyms

EEF 2 antibody

Eef2 antibody

EF-2 antibody

EF2 antibody

EF2_HUMAN antibody

Elongation factor 2 antibody

Eukaryotic translation elongation factor 2 antibody

Polypeptidyl tRNA translocase antibody

SCA26 antibody

Images

  • <span style="font-weight: bold;">☑ Cell treatment (CT)</span><br /><br />Western blot analysis of Phospho-EEF2 (T56) on different lysates with Rabbit anti-Phospho-EEF2 (T56) antibody (<a href="/products/HA751075" style="font-weight: bold;text-decoration: underline;">HA751075</a>) at 1/2,000 dilution.<br /><br />Lane 1: HeLa cell lysate<br />Lane 2: HeLa starved for 24 hours then add 10μM forskolin for 1 hour cell lysate<br />Lane 3: NIH/3T3 cell lysate<br />Lane 4: NIH/3T3 starved for 24 hours then add 10μM forskolin for 1 hour cell lysate<br />Lane 5: PC-12 cell lysate<br />Lane 6: PC-12 treated with 20μM forskolin for 1 hour then add 100nM Calyculin A for 30 minutes cell lysate<br />Lane 7: HepG2 cell lysate<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 95 kDa<br />Observed band size: 95 kDa<br /><br />Exposure time: 3 minutes; ECL: K1802;<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/HA751075" style="font-weight: bold;text-decoration: underline;">HA751075</a>) at 1/2,000 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.

    ☑ Cell treatment (CT)

    Western blot analysis of Phospho-EEF2 (T56) on different lysates with Rabbit anti-Phospho-EEF2 (T56) antibody (HA751075) at 1/2,000 dilution.

    Lane 1: HeLa cell lysate
    Lane 2: HeLa starved for 24 hours then add 10μM forskolin for 1 hour cell lysate
    Lane 3: NIH/3T3 cell lysate
    Lane 4: NIH/3T3 starved for 24 hours then add 10μM forskolin for 1 hour cell lysate
    Lane 5: PC-12 cell lysate
    Lane 6: PC-12 treated with 20μM forskolin for 1 hour then add 100nM Calyculin A for 30 minutes cell lysate
    Lane 7: HepG2 cell lysate

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 95 kDa
    Observed band size: 95 kDa

    Exposure time: 3 minutes; ECL: K1802;

    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 (HA751075) at 1/2,000 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.

  • Immunohistochemical analysis of paraffin-embedded rat liver tissue with Rabbit anti-Phospho-EEF2 (T56) antibody (<a href="/products/HA751075" style="font-weight: bold;text-decoration: underline;">HA751075</a>) at 1/500 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/HA751075" style="font-weight: bold;text-decoration: underline;">HA751075</a>) at 1/500 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 liver tissue with Rabbit anti-Phospho-EEF2 (T56) antibody (HA751075) at 1/500 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 (HA751075) at 1/500 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.

  • Immunocytochemistry analysis of HeLa cells starved for 24 hours then add 10μM forskolin for 1 hour labeling Phospho-EEF2 (T56) with Rabbit anti-Phospho-EEF2 (T56) antibody (<a href="/products/HA751075" style="font-weight: bold;text-decoration: underline;">HA751075</a>) at 1/100 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-EEF2 (T56) antibody (<a href="/products/HA751075" style="font-weight: bold;text-decoration: underline;">HA751075</a>) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/M1305-2" style="font-weight: bold;text-decoration: underline;">M1305-2</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    Immunocytochemistry analysis of HeLa cells starved for 24 hours then add 10μM forskolin for 1 hour labeling Phospho-EEF2 (T56) with Rabbit anti-Phospho-EEF2 (T56) antibody (HA751075) at 1/100 dilution.

    Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-EEF2 (T56) antibody (HA751075) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • <span style="font-weight: bold;">☑ Cell treatment (CT)</span><br /><br />Immunocytochemistry analysis of PC-12 cells treated with 20μM forskolin for 1 hour then add 100nM Calyculin A for 30 minutes labeling Phospho-EEF2 (T56) with Rabbit anti-Phospho-EEF2 (T56) antibody (<a href="/products/HA751075" style="font-weight: bold;text-decoration: underline;">HA751075</a>) at 1/100 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-EEF2 (T56) antibody (<a href="/products/HA751075" style="font-weight: bold;text-decoration: underline;">HA751075</a>) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/M1305-2" style="font-weight: bold;text-decoration: underline;">M1305-2</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    ☑ Cell treatment (CT)

    Immunocytochemistry analysis of PC-12 cells treated with 20μM forskolin for 1 hour then add 100nM Calyculin A for 30 minutes labeling Phospho-EEF2 (T56) with Rabbit anti-Phospho-EEF2 (T56) antibody (HA751075) at 1/100 dilution.

    Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-EEF2 (T56) antibody (HA751075) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

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