Human meningioma samples were obtained from the Brigham and Women’s Hospital Department of Pathology, under institutional IRB approval. Protein from meningioma samples was extracted using RIPA buffer (Cell Signaling, Danvers, USA). Cultured meningioma cells with or without treatment were lysed using cell lysis buffer (Cell Signaling, Danvers, USA). 20 μg of protein from each sample was resolved by 4–20% SDS-PAGE (Bio-Rad), before transfer to a PVDF membrane for overnight incubation with primary antibodies at 4 °C. Primary antibodies were diluted at 1:1000 for OGN (Abcam, Cambridge, USA), cyclinD1, cyclinA2, cyclinB1, GAPDH, p-AKT(Ser473), p-mTOR (Ser2448), p-4E-BP1(Ser65), p-eIF4B (Ser422), p-eEF2K (Ser366), p-ULK(Ser757), NF2, LC3IIB (Cell Signaling, Danvers, USA). Membranes were then incubated with horseradish peroxidase-conjugated goat anti-rabbit or goat anti-mouse IgG (Cell Signaling, Danvers, USA) at 1:2000. Blots were visualized by an ECL system (GE Healthcare, Buckinghamshire, UK) and protein quantified by densitometry analyses using ImageJ.
P eif4b
Manufactured by Cell Signaling Technology
Sourced in United States
P-eIF4B is a phospho-specific antibody that recognizes the phosphorylated form of the eukaryotic translation initiation factor 4B (eIF4B) protein. eIF4B is involved in the regulation of protein synthesis initiation.
Automatically generated - may contain errors
Lab products found in correlation
P mtor, by Cell Signaling Technology (3 mentions)
P 4ebp1, by Cell Signaling Technology (3 mentions)
P eif4e, by Cell Signaling Technology (3 mentions)
4e bp1, by Cell Signaling Technology (3 mentions)
P akt, by Cell Signaling Technology (2 mentions)
Actin, by Santa Cruz Biotechnology (2 mentions)
Eif4g, by Cell Signaling Technology (2 mentions)
Eif4e, by Cell Signaling Technology (2 mentions)
Eif4a, by Cell Signaling Technology (2 mentions)
P eif4g, by Cell Signaling Technology (2 mentions)
Eif4b, by Cell Signaling Technology (2 mentions)
Horseradish peroxidase conjugated goat anti rabbit or goat anti mouse igg, by Cell Signaling Technology (1 mentions)
Cell lysis buffer, by Cell Signaling Technology (1 mentions)
Sds page, by Bio-Rad (1 mentions)
Ecl system, by GE Healthcare (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
Ripa buffer, by Cell Signaling Technology (1 mentions)
Rabbit anti human β actin antibody, by Merck Group (1 mentions)
Z devd fmk, by Merck Group (1 mentions)
P p70s6k, by Cell Signaling Technology (1 mentions)
5 protocols using p eif4b
1
Meningioma Protein Expression Analysis
2
Purification of 13-Acetoxysarcocrassolide from Soft Coral
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
13-Acetoxysarcocrassolide was purified after extracting it from the cultured Formosa soft coral S. crassocaule, according to the method described by others (Duh et al. 2000 (link)). Dimethyl sulfoxide (DMSO), 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT), Z-DEVD-FMK (caspase-3 inhibitor), Z-VAD-FMK (a cell-permeable pan-caspase inhibitor), a protease inhibitor cocktail, and rabbit anti-human β-actin antibodies were purchased from Sigma (St. Louis, MO, USA). Antibodies against pro-caspase 9, cleaved-caspase 9, pro-caspase 3, cleaved-caspase 3, cleaved-PARP-1, Bax, Bad, Bcl-xL, Mcl-1, p-Bad, PARP-1, Bcl-2, PI3K, p-PI3K, AKT, p-AKT, mTOR, p-mTOR, p70S6K, p-p70S6K, 4EBP1, p-4EBP1, p-S6, p-eIF4B, and p-eIF4E, and goat anti-rabbit and horseradish peroxidase-conjugated immunoglobulin G were obtained from Cell Signalling Technology (Danvers, MA, USA). Dulbecco’s modified Eagle’s medium (DMEM) and foetal bovine serum (FBS) were obtained from Biowest (Nuaillé, France).
3
Western Blotting Protein Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting analyses were performed as described previously (Marabita et al., 2016 (link)). Antibodies for P-Akt, Akt, eIF4E, P-eIF4E, P-eIF4B, eIF4B, eIF4A, eIF4H, eIF4G, P-eIF4G, 4E-BP1, P-S6, S6 were taken from Cell Signaling. Actin was from Santa Cruz, and puromycin from Millipore. All quantifications of the western blots were done on at least four different blots for each protein, in each condition. Differences between groups were assessed using Student's t-test. Significance was defined as a value of P < 0.05 (95% confidence).
4
Modulation of THP-1 and MM-6 Differentiation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Differentiation of THP-1 and MM-6 cells was performed using Phorbol 12-myristate 13-acetate (PMA; Merck Millipore) or 1-α, 25-dihydroxyvitamin D3 (VitD3; Sigma Aldrich). For the inhibition of kinases and proteases, the following inhibitors were applied: S6K1 inhibitor DG2 (S6K-I), PKR inhibitor (PKR-I; 6,8-Dihydro-8-1H-imidazol-5-ylmethylene)-7H-pyrrolo[2,3-g]benzothiazol-7-one), ALLN (N-[N-(N-Acetyl-L-leucyl)-L-leucyl]-L-norleucine), MG-132, Calpain-Inhibitor V (Merck Millipore), rapamycin (mTOR-I; Sigma Aldrich), UO126 (MEK-I; Cell Signaling, Danvers, USA), BI-D1870 (RSK-I; Axon Medchem BV, Groningen, Netherland), and Cathepsin-Inhibitor I (Applichem). For Western Blotting, specific antibodies directed against RSK, phosphorylated (p-)RSK (T573), ERK1/2, p-ERK1/2 (T202/Y204), rpS6, p-rpS6 (S335/236), eIF4B, p-eIF4B (S422), eIF4A, eIF2α, p-eIF2α (S51), PKR (Cell Signaling), p-PKR (T446; Abcam, Cambridge, UK), V5 Tag (Invitrogen), C/EBPβ, Calpastatin, Histon H4 (Santa Cruz), Actin, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), or TATA box-binding protein (TBP; Sigma Aldrich) were used in combination with secondary horseradish peroxidase (HRP-)coupled antibodies (Vector Laboratories, Peterborough, UK). All media and reagents were of the best available grade and routinely tested for endotoxins with the Limulus Amoebocyte Lysate assay (Lonza).
5
Protein Expression Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Primary antibodies from Cell Signaling Technology were used to analyze the expression of important proteins, Bcl-2 (catalog #4223S), Bax (catalog #2772S), Bcl-xL (catalog #2764S), eIF4e (catalog #9742S), peIF4e (catalog #9741S), 4EBP1 (catalog #9452S), p4EBP1 (catalog #2855S), mTOR (catalog #4517S), p-mTOR (catalog #5536S), eIF4B (catalog #3592S), peIF4B (catalog #3591S), eIF4G (catalog #2498S), peIF4G (catalog #2441S), p18 (catalog # 2896S) and p21 (catalog #2947S). Cleaved PARP (sc-8007) and Cyclin D1 (sc-753) antibodies were obtained from Santa Cruz Biotechnology (Dallas, TX, USA). β-actin antibody was obtained from Sigma (catalog #A2228; St. Louis, MO, USA). The horseradish peroxidase conjugated rabbit (catalog #4011) and mouse (catalog #4021) secondary antibodies were procured from Promega (Madison, WI, USA).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!