- 44-530G - Invitrogen Antibodies EIF2B5 antibody

Submitted references Proximal tubular epithelial insulin receptor mediates high-fat diet-induced kidney injury.

Lee HJ, Mariappan MM, Norton L, Bakewell T, Feliers D, Oh SB, Donati A, Rubannelsonkumar CS, Venkatachalam MA, Harris SE, Rubera I, Tauc M, Ghosh Choudhury G, Kahn CR, Sharma K, DeFronzo RA, Kasinath BS
JCI insight 2021 Feb 8;6(3)

Pharmacological inhibition of GSK-3 in a guinea pig model of LPS-induced pulmonary inflammation: II. Effects on skeletal muscle atrophy.

Verhees KJ, Pansters NA, Baarsma HA, Remels AH, Haegens A, de Theije CC, Schols AM, Gosens R, Langen RC
Respiratory research 2013 Nov 1;14(1):117

Glycogen synthase kinase-3β is required for the induction of skeletal muscle atrophy.

Verhees KJ, Schols AM, Kelders MC, Op den Kamp CM, van der Velden JL, Langen RC
American journal of physiology. Cell physiology 2011 Nov;301(5):C995-C1007

Nutrient provision increases signalling and protein synthesis in human skeletal muscle after repeated sprints.

Coffey VG, Moore DR, Burd NA, Rerecich T, Stellingwerff T, Garnham AP, Phillips SM, Hawley JA
European journal of applied physiology 2011 Jul;111(7):1473-83

Pulmonary artery smooth muscle hypertrophy: roles of glycogen synthase kinase-3beta and p70 ribosomal S6 kinase.

Deng H, Hershenson MB, Lei J, Anyanwu AC, Pinsky DJ, Bentley JK
American journal of physiology. Lung cellular and molecular physiology 2010 Jun;298(6):L793-803

Ectopic expression of eIF2Bepsilon in rat skeletal muscle rescues the sepsis-induced reduction in guanine nucleotide exchange activity and protein synthesis.

Tuckow AP, Vary TC, Kimball SR, Jefferson LS
American journal of physiology. Endocrinology and metabolism 2010 Aug;299(2):E241-8

Resistance exercise increases muscle protein synthesis and translation of eukaryotic initiation factor 2Bepsilon mRNA in a mammalian target of rapamycin-dependent manner.

Kubica N, Bolster DR, Farrell PA, Kimball SR, Jefferson LS
The Journal of biological chemistry 2005 Mar 4;280(9):7570-80

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Peptide Competition. Recombinant eIF2Bepsilon treated with lambda phosphatase (1) or GSK-3beta (2-5) were added to background extracts and resolved by SDS-PAGE on a 10% Tris-glycine gel and transferred to PVDF. The membrane was blocked with a 5% BSA-TBST buffer overnight at 4°C, then incubated with the eIF2Bepsilon (pS539) antibody for two hours at room temperature in a 1% BSA-TBST buffer, following prior incubation with: no peptide (1, 2), the non-phosphopeptide corresponding to the phosphopeptide immunogen (3), a generic phosphoserine-containing peptide (4), or the phosphopeptide immunogen (5). After washing, the membrane was incubated with goat F (ab’)2 anti-rabbit IgG alkaline phosphatase (Product # ALI4405) and signals were detected using the Tropix WesternStar™ method. The data show that only the peptide corresponding to eIF2Bepsilon (pS539) blocks the antibody signal, demonstrating the specificity of the antibody. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody is phospho-specific. The recombinant eIF2Bepsilon was kindly provided by Dr. Scot R. Kimball, Penn State University.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on whole cell extracts (30 µg lysate) of L6 (Lane 1), L6 differentiated to myotubes (Lane 2), L6 treated with Insulin (10nM for 10 min) (Lane 3), C2C12 (Lane 4) and C2C12 treated with Insulin (10nM for 10 min) ( (Lane 5). The blot was probed with Anti-Phospho-eIF2b epsilon (Ser539) Polyclonal Antibody (Product # 44-530G, 1:2000 dilution) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). A 80 kDa band corresponding to Phospho-eIF2b epsilon was observed across the cell lines tested and reduced upon treatment and in differentiated cell line.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 Muscle protein turnover signaling is not affected following chronic LPS-treatment and GSK-3 inhibition. Protein synthesis and protein degradation-related signaling molecules were determined in whole muscle homogenates of the extensor digitorum longus (EDL) muscles by Western blot analysis of guinea pigs that were treated intranasally with LPS or SB216763 for 12 weeks. Representative immunoblots depict protein levels of (A) phospho-Akt (p-Akt), total Akt, phospho-GSK-3beta (p-GSK-3beta), total GSK-3beta, phospho-eukaryotic initiation factor 2B (p-eIF2B), GAPDH, (B) phospho-mammalian target of rapamycin (p-mTOR), total mTOR, phospho-p70S6K (p-p70S6K), total p70S6K, phospho-S6 (p-S6), phospho-4E-BP1 (p-4E-BP1), total 4E-BP1, GAPDH, (C) phospho-FoXO1 (p-FoXO1), total FoXO1, phospho-FoXO3a (p-FoXO3a), total FoXO3a and GAPDH. The accompanying bar graphs show the densitometric analysis results (means +- SEM, n = 6), represented as a percentage of the vc control group corrected for GAPDH. All data is shown as a ratio of phospho- to total protein for each target (except p-eIF2B, p-S6 and (p-) 4E-BP1). *p < 0.05 compared with the vc control group. NS: not significant.

44-530G

Antibody data