U87 and T98G cells were lysed in radioimmunoprecipitation assay buffer [150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulfate (SDS), 50 mM Tris, pH 8.0, 5.0 mM ethylenediaminetetraacetic acid, pH 8.0, 0.5 mM dithiothreitol and 1 mM phenylmethylsulfonyl fluoride]. Protein concentrations were determined using the bicinchoninic acid method (Thermo Scientific, Rockford, IL, USA). Protein lysates were separated by SDS-polyacrylamide gel electrophoresis and transferred onto polyvinylidene difluoride membranes (Millipore, Billerica, MA, USA) by electroblotting. Primary antibodies for immunodetection were anti-CEP55 (Santa Cruz), anti-GLUT1 (Santa Cruz), anti-p-AktS473 (Santa Cruz), anti-p-AktT308(Santa Cruz), anti-Akt (Santa Cruz), anti-p-mTOR (Santa Cruz), anti-mTOR (Santa Cruz), anti-BAD (Santa Cruz), anti-caspase-9 (Santa Cruz), anti-GSK3-β (Santa Cruz), anti-p27 (Abcam) and anti-GAPDH (Santa Cruz). Subsequent to being incubated with Horseradish peroxidase (HRP) conjugated anti-rabbit or anti-mouse secondary antibodies (1: 10000, Santa Cruz) for 1 h, the immune complexes were detected using the enhanced chemiluminescence method.
Anti p mtor
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Anti-p-mTOR is a primary antibody that specifically recognizes the phosphorylated form of the mammalian target of rapamycin (mTOR) protein. mTOR is a serine/threonine protein kinase that plays a crucial role in cellular growth, proliferation, and metabolism.
Automatically generated - may contain errors
Lab products found in correlation
Anti mtor, by Santa Cruz Biotechnology (7 mentions)
Anti akt, by Santa Cruz Biotechnology (4 mentions)
Anti β actin, by Santa Cruz Biotechnology (4 mentions)
Anti akt, by Cell Signaling Technology (3 mentions)
Anti p akt, by Cell Signaling Technology (3 mentions)
Anti p akt, by Santa Cruz Biotechnology (3 mentions)
Enhanced chemiluminescence detection kit, by Thermo Fisher Scientific (3 mentions)
Anti parp1, by Proteintech (3 mentions)
Anti ire1α, by Proteintech (3 mentions)
Anti atf4, by Proteintech (3 mentions)
Anti grp78, by Proteintech (3 mentions)
Anti par, by Bio-Techne (3 mentions)
Anti kitl, by Biorbyt (3 mentions)
Anti p ire1α, by Biorbyt (3 mentions)
Anti p perk, by Biorbyt (3 mentions)
Anti ha tag, by Yeasen (3 mentions)
Anti flag tag, by Yeasen (3 mentions)
Tanon 4500 gel imaging system, by Thermo Fisher Scientific (3 mentions)
Anti perk, by Proteintech (3 mentions)
Anti gapdh, by Abcam (3 mentions)
16 protocols using anti p mtor
1
Protein Expression Analysis in Glioblastoma Cells
2
Vitamin D Analog TX 527 Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Vitamin D analog TX 527 [19-nor-14,20-bisepi-23-yne-1,25(OH)2D3] was originally synthesized by M. Vandewalle and P. De Clercq (University of Ghent, Ghent, Belgium) and provided by Théramex (Monaco). Immobilon P (polyvinylidenedifluoride; PVDF) membranes, 1α,25(OH)2D3, the antibiotic G418 and LY294002 were from Sigma-Aldrich (St. Louis, MO, USA). Puromycin supplied by Invivogen (San Diego, CA, USA). The antibodies used were rabbit monoclonal anti-p-Akt, anti-Akt, anti-MEKα (Cell Signaling Technology, Danvers, MA, USA) and anti-Tubulin (Thermo Fisher Scientific Inc., Waltham, MA, USA); mouse monoclonal anti-LC3, anti-BECN1, anti-p-mTOR, and anti-mTOR, goat polyclonal Lamin B and secondary antibodies anti-rabbit, anti-mouse and anti-goat (Santa Cruz Biotechnology, Santa Cruz, CA, USA). Roche Applied Science supplied high Pure RNA Isolation Kit (Indianapolis, IN, USA). High Capacity cDNA RT and SYBR Green PCR Master Mix reagent (Applied Biosystems) were acquired from Thermofisher (Thermofisher, Buenos Aires, AR). GAPDH primer was from InvitrogenTM (Invitrogen/Thermo Fisher Scientific Inc., Waltham, MA, USA) and BECN1 primer was obtained by Eurogentec (Serain, Belgium). The inhibitor of flux autophagy, Chloroquine, was kindly provided by Dr. Daniel Grasso (IBIMOL Universidad de Buenos Aires – CONICET).
3
Inhibition of PAK4, NAMPT, and mTOR Signaling
4
Protein Expression Analysis in Midbrain
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The middle brain from each experimental group was removed and homogenized in RIPA lysis buffer containing protease and phosphatase cocktail (Sigma Aldrich, Cat No. #P8340, Cat No. P-0044). The supernatant was collected, and protein quantification was performed by the BCA method. An equal amount (50μg) of protein was loaded and separated by SDS-PAGE before being blotted or transferred onto nitrocellulose membrane and blocked for 2 h in 3% bovine serum albumin (BSA) in TBST at room temperature. After that membrane was incubated with primary antibodies (Anti-Atg5, 12994P), Anti-Atg7(8558T), Anti-Beclin (3495S), LC3B (12741S), Anti-mTOR(2983P) Cell signalling technology, USA) (1:2000) Anti-p62 (SC25575), Anti-β-Actin (SC47778), anti-P-mTOR (SC293133), Anti-HO1, Anti-α-Synuclein (SC53955), and Anti NRF2(12721S) (Santa Cruz, USA) (1:1000), at 4°C for overnight. Following washing with TBST, membranes were incubated with horseradish peroxidase (HRP) conjugated anti-mouse (Cat No SC516102) and anti-Rabbit (Cat no SC2357) secondary antibodies (Santa Cruz, USA) (1:15000), and these primary-secondary antibodies complex chemiluminescence was observed by Fusion-FX chemiluminescence imager (Vilber Lourmat, Germany). Later, Image-J software was used to calculate the relative band densities [20 (link)].
5
Con A-Induced Hepatitis Model
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Con A and dimethyl sulfoxide were purchased from Sigma-Aldrich (St Louis, MO, USA) and stored at 4°C. PAG was purchased from Sigma-Aldrich, and the RNA polymerase chain reaction (PCR) kit was obtained from Takara Biotechnology (Dalian, People’s Republic of China). Fetal bovine serum and Dulbecco’s Modified Eagle’s Medium were obtained from Thermo Fisher Scientific (Waltham, MA, USA). LC3-2 and Beclin-1 were purchased from Abcam (Cambridge, UK). The antibodies used for immunoblotting and immunohistochemical staining were anti-tumor necrosis factor (TNF)-α, anti-interleukin (IL)-6, anti-mTOR, anti-p-mTOR, anti-p-AKT, and anti-AKT (Santa Cruz Biotechnology, Dallas, TX, USA). Con A was dissolved in pyrogen-free physiological saline and intravenously injected at a dose of 20 mg/kg body weight to induce hepatitis as previously described.1 (link) NaHS was dissolved in saline and intravenously injected at a dose of 14 μmol/kg body weight. PAG was dissolved in pyrogen-free physiological saline and intraperitoneally injected at a dose of 50 mg/kg body weight.
6
In Situ Immunofluorescence Analysis of mTOR Signaling in T Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
In situ expression of p-mTOR/mTOR in T cells of treated mice was detected by immunofluorescence. Mouse spleen tissues were deparaffinized, permeabilized in 0.1% Triton X-100 in phosphate buffered saline (PBS) for 30 min, and blocked in PBS with 1% bovine serum albumin (BSA) for one hour. Samples were then incubated with a mixture of rabbit mAb Anti-mTOR (1:100, Catalog #: 55306F, ABMART, Shanghai China) and mouse mAb Anti-p-mTOR (1:100, Catalog #: 293133, Santa Cruz Biotechnology, Santa Cruz, CA, USA) and PE Anti-Mouse IL-17A antibody (1:100, Catalog #: 1995433, BD Biosciences, Franklin Lakes, NJ, USA) overnight at 4 °C. After rinsing with PBS. The samples were incubated with a solution containing the fluorescent secondary antibodies Alexa Fluor 647 AffiniPure Goat Anti-Mouse immunoglobulin (Ig) G (Catalog #: 115-605-205, Jackson ImmunoResearch Inc., West Grove, PA, USA) or Alexa Fluor 594 AffiniPure Goat Anti-Rabbit IgG (Catalog #: 115-585-003, Jackson ImmunoResearch Inc., West Grove, PA, USA) for 2 h. After washing with PBS, DAPI was applied for 10 min to stain the nuclei, and the slides were sealed with a drop of anti-fluorescence quenching mounting solution. A Leica TCS SP8 laser scanning confocal microscope and ImageJ software were employed to capture and quantify the resulting images.
7
Comprehensive Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total ovary lysates were separated on SDS-PAGE. After electrophoresis, the protein was transferred to PVDF membrane. The membranes were blocked with 5% skimmed milk and incubated with anti-GRP78 (1:1000, #11587–1-AP, Proteintech, Hubei, China), anti-PARP1 (1:4000, #66520-1-lg, Proteintech), anti-PAR (1:1000, #4335-MC-100, Bio-techne, Minneapolis, USA), anti-mTOR (1:1000, #20657-1-AP, Proteintech), anti-p-mTOR (1:800, #sc-293133, Santa cruz, Dallas, Texas, USA), anti-KitL (1:1000, #bs-0545R, Biorbyt, Hubei, China), anti-p-IRE1α (1:800#, bs-16698R, Biorbyt), anti-IRE1α (1:1000, #27528-1-AP, Proteintech), anti-ATF4 (1:1000, #10835-1-AP, Proteintech), anti-p-PERK (1:800, #Orb336657, Biorbyt), anti-PERK (1:1000, #24390-1-AP, Proteintech), anti-HA-tag (1:1000, #30701ES60, YEASEN), anti-FLAG-tag (1:1000, #30501ES60, YEASEN) and anti-GAPDH (1:4000, #ab8245, Abcam, Cambridge, UK) antibodies overnight at 4 °C. After TBST washing, the membranes were treated with the corresponding peroxidase-conjugated secondary antibody, and then the signal was detected with an Enhanced Chemiluminescence Detection Kit (#32106, Thermo Scientific, Waltham, MA, USA) on Tanon 4500 gel imaging system (Shanghai, China).
8
Comprehensive Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total ovary lysates were separated on SDS-PAGE. After electrophoresis, the protein was transferred to PVDF membrane. The membranes were blocked with 5% skimmed milk and incubated with anti-GRP78 (1:1000, #11587–1-AP, Proteintech, Hubei, China), anti-PARP1 (1:4000, #66520-1-lg, Proteintech), anti-PAR (1:1000, #4335-MC-100, Bio-techne, Minneapolis, USA), anti-mTOR (1:1000, #20657-1-AP, Proteintech), anti-p-mTOR (1:800, #sc-293133, Santa cruz, Dallas, Texas, USA), anti-KitL (1:1000, #bs-0545R, Biorbyt, Hubei, China), anti-p-IRE1α (1:800#, bs-16698R, Biorbyt), anti-IRE1α (1:1000, #27528-1-AP, Proteintech), anti-ATF4 (1:1000, #10835-1-AP, Proteintech), anti-p-PERK (1:800, #Orb336657, Biorbyt), anti-PERK (1:1000, #24390-1-AP, Proteintech), anti-HA-tag (1:1000, #30701ES60, YEASEN), anti-FLAG-tag (1:1000, #30501ES60, YEASEN) and anti-GAPDH (1:4000, #ab8245, Abcam, Cambridge, UK) antibodies overnight at 4 °C. After TBST washing, the membranes were treated with the corresponding peroxidase-conjugated secondary antibody, and then the signal was detected with an Enhanced Chemiluminescence Detection Kit (#32106, Thermo Scientific, Waltham, MA, USA) on Tanon 4500 gel imaging system (Shanghai, China).
9
Comprehensive Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total ovary lysates were separated on SDS-PAGE. After electrophoresis, the protein was transferred to PVDF membrane. The membranes were blocked with 5% skimmed milk and incubated with anti-GRP78 (1:1000, #11587–1-AP, Proteintech, Hubei, China), anti-PARP1 (1:4000, #66520-1-lg, Proteintech), anti-PAR (1:1000, #4335-MC-100, Bio-techne, Minneapolis, USA), anti-mTOR (1:1000, #20657-1-AP, Proteintech), anti-p-mTOR (1:800, #sc-293133, Santa cruz, Dallas, Texas, USA), anti-KitL (1:1000, #bs-0545R, Biorbyt, Hubei, China), anti-p-IRE1α (1:800#, bs-16698R, Biorbyt), anti-IRE1α (1:1000, #27528-1-AP, Proteintech), anti-ATF4 (1:1000, #10835-1-AP, Proteintech), anti-p-PERK (1:800, #Orb336657, Biorbyt), anti-PERK (1:1000, #24390-1-AP, Proteintech), anti-HA-tag (1:1000, #30701ES60, YEASEN), anti-FLAG-tag (1:1000, #30501ES60, YEASEN) and anti-GAPDH (1:4000, #ab8245, Abcam, Cambridge, UK) antibodies overnight at 4 °C. After TBST washing, the membranes were treated with the corresponding peroxidase-conjugated secondary antibody, and then the signal was detected with an Enhanced Chemiluminescence Detection Kit (#32106, Thermo Scientific, Waltham, MA, USA) on Tanon 4500 gel imaging system (Shanghai, China).
10
Adipogenesis Regulation via Autophagy
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Icariin, L-asparagine (Asn), 3-methyladenine (3-MA), Oil Red O, MTT assay kit, uranyl acetate/lead citrate, and MDC were purchased from Sigma-Aldrich, Inc. (St. Louis, MO, USA). Dulbecco's modified Eagle's medium (DMEM), fetal bovine serum (FBS), penicillin, adipocyte differentiation medium, and gentamycin were purchased from Hyclone Laboratories, Inc. (Logan, UT, USA). The Annexin V-FITC apoptosis detection kit was purchased from BD Biosciences (Franklin Lakes, NJ, USA). Anti-mTOR, anti-p-mTOR, anti-beclin-1, anti-AMPK, anti-p-AMPK, anti-p62, anti-LC3, and anti-β-actin antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, 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!