- PA5-17794 - Invitrogen Antibodies EEF2 antibody

Submitted references α1AMP-Activated Protein Kinase Protects against Lipopolysaccharide-Induced Endothelial Barrier Disruption via Junctional Reinforcement and Activation of the p38 MAPK/HSP27 Pathway.

Angé M, Castanares-Zapatero D, De Poortere J, Dufeys C, Courtoy GE, Bouzin C, Quarck R, Bertrand L, Beauloye C, Horman S
International journal of molecular sciences 2020 Aug 4;21(15)

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of EEF2 using a polyclonal antibody (Product # PA5-17794).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of eEF2 in HeLa cells using an EEF2 polyclonal antibody (Product # PA5-17794) (green). Actin filaments are labeled with a fluorescent red phalloidin. DNA is labeled using a fluorescent blue dye.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 1 Basal alpha1AMPK regulates VE-Cad, ZO-1, and Cx43 expression. ( a ) Schematic representation of the experimental design, in which HMECs were transfected with control non-targeting siRNA or alpha1AMPK siRNA (50 nM) for 48 h. Then, they were treated with 991 (1 muM) or DMSO for one hour and subsequently exposed or not exposed to lipopolysaccharide (LPS, 50 muM) for six hours; ( b , c ) Human dermal microvascular endothelial cells (HMECs) were treated according to the protocol detailed in ( a ). Cell lysates were submitted to Western blot analysis and probed with alpha1AMPK, phospho-AMPK (Thr172), phospho-ACC (Ser79), VE-Cad, ZO-1, and Cx43 antibodies (Abs). Anti-eukaryotic elongation factor 2 (eEF2) was used as a loading control. Representative western blots; ( b ) and quantifications ( c ) are shown. Data are expressed as mean +- standard deviation (SD) (three biological replicates for each condition). # p < 0.05 is relative to corresponding untreated HMECs and * p < 0.05 is relative to cells transfected with the scrambled siRNA. The data underwent two-way analysis of variance (ANOVA).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 Cx43 deficiency is associated with decreased VE-Cad and ZO-1 expression. ( a - e ) HMECs were transfected with control non-targeting siRNA or Cx43 siRNA (50 nm) for 48 h. ( a - c ) Cell lysates were submitted to Western blot analysis and probed with Cx43 ( a ), VE-Cad ( b ), and ZO-1 ( c ) Abs. eEF2 was used as a loading control. Representative Western blots ( top panel) and quantifications ( bottom panel) are shown; ( d , e ) Cx43, VE-Cad, and ZO-1 immunostainings. Nuclei are stained with DAPI. Scale bar, 50 mum. Representative images ( d ) and quantifications ( e ) are shown. Data are expressed as mean +- SD (three biological replicates for each condition). * p < 0.05 is relative to cells transfected with the scrambled siRNA. Analyses were performed using Student's t -test.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 AMPK Activation Leads to Increased HSP27 Phosphorylation, via a p38 MAPK-Dependent Way. ( a , b ) HMECs were transfected with control non-targeting siRNA or alpha1AMPK siRNA (50 nM) for 48 h and then treated with DMSO or 991 (1 muM) for one hour. They were then exposed to LPS (50 muM) for six hours; ( c , d ) HMECs were preincubated with DMSO or with the p38 MAPK inhibitor SB203580 (10 muM) for 15 min. Then, they were treated with 991 (1 muM) or DMSO for one hour and subsequently, either exposed or not exposed to LPS (50 muM) for six hours. ( a - d ) Cell lysates were submitted to western blot analysis and probed with total HSP27, total p38 MAPK, phospho-HSP27 and phospho-p38 MAPK Abs. eEF2 was used as a loading control. Representative Western blots ( a , c ) and quantification of HSP27 (Ser82) phosphorylation ( b , d ) are shown. Data are expressed as means +- SD (three biological replicates for each condition). # p < 0.05 is relative to corresponding non-treated HMECs, $ p < 0.05 is relative to corresponding LPS-treated HMECs, and * p < 0.05 is relative to cells transfected with non-targeting siRNA or that were treated with the vehicle. The data underwent two-way ANOVA.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 7 AMPK Regulates Actin Organization Independently of Rac1. ( a , b ) HMECs were treated with saline or the Rac1 inhibitor NSC23766 (100muM) for 15 min before adding DMSO or 991 (1muM) for one hour. Then, they were exposed to LPS (50muM) for six hours. ( a ) Lysates were engaged in G-LISA assays to precisely monitor Rac1 activity. Quantification represents the differential signal, compared to basal, in fold of control; ( b ) Representative Western blots and quantification of HSP27 (Ser82) phosphorylation. eEF2 was used as a loading control. Data are expressed as means +- SD (three biological replicates for each condition). # p < 0.05 is relative to corresponding non-treated HMECs. * p < 0.05 is relative to cells treated with the vehicle. The data underwent two-way ANOVA.

PA5-17794

Antibody data