Total protein was extracted from peritoneal macrophages using Cell Lysis Buffer (Cell Signaling Technology Inc., Danvers, MA) according to the manufacturer’s instructions. Protein concentrations were determined with a bicinchoninic acid (BCA) assay (Pierce BCA Protein Assay Kit, Thermo Fish Scientific Inc.). Proteins were separated on 4–20% gradient SDS/polyacrylamide gels and blotted onto polyvinylidene difluoride (PVDF) membranes. The following antibodies were used as primary antibodies: anti-PERK (C33E10, 1:2000), anti-BiP (1:2000), anti-CHOP (D46F1, 1:2000) (Cell Signaling Technology Inc.), and anti-β-actin (AC-15, 1:10,000, Sigma–Aldrich). Anti-Rabbit IgG, HRP-linked whole Ab donkey (GE Healthcare) was used as secondary antibody (1:5000). Chemiluminescence images were acquire with an Image Quant LAS-4010 (GE Healthcare). Densitometric analyses were performed using an Image Quant TL (GE Healthcare).
Anti perk c33e10
Manufactured by Cell Signaling Technology
Sourced in Panama
The Anti-PERK (C33E10) is a primary antibody used for the detection of PERK protein in various applications, such as Western blotting, immunoprecipitation, and immunohistochemistry. PERK is an endoplasmic reticulum (ER) transmembrane protein that plays a key role in the unfolded protein response (UPR) pathway.
Automatically generated - may contain errors
Lab products found in correlation
Anti p63 4a4, by Santa Cruz Biotechnology (2 mentions)
Gapdh, by Merck Group (2 mentions)
P124 anti dsg1 extracellular domain, by Progen Biotechnik (2 mentions)
27b2 anti dsg1 cytodomain, by Thermo Fisher Scientific (2 mentions)
U100 anti dsc1, by Progen Biotechnik (2 mentions)
Hecd1 anti e cadherin, by Takara Bio (2 mentions)
Anti erk1 2, by Promega (2 mentions)
Imagequant tl, by GE Healthcare (1 mentions)
Cell lysis buffer, by Cell Signaling Technology (1 mentions)
Imagequant las 4010, by GE Healthcare (1 mentions)
Anti β actin ac 15, by Merck Group (1 mentions)
Anti bip, by Cell Signaling Technology (1 mentions)
Anti rabbit igg hrp linked whole ab donkey, by GE Healthcare (1 mentions)
Protease inhibitor cocktail, by Nacalai Tesque (1 mentions)
Image studio software, by LI COR (1 mentions)
Blocking one p, by Nacalai Tesque (1 mentions)
Westernsure ecl substrate, by LI COR (1 mentions)
Ez capture 2, by ATTO Corporation (1 mentions)
Anti flag m2, by Merck Group (1 mentions)
Anti gcn2, by Cell Signaling Technology (1 mentions)
4 protocols using anti perk c33e10
1
Western Blot Analysis of ER Stress Markers
2
Immunoblot Analysis of Stress Kinases
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Cells were lysed in TNT buffer (50 mM Tris, pH 7.5, 150 mM NaCl, 10% glycerol, 1% Triton X-100) with protease inhibitor cocktail (Nacalai Tesque), phosphatase inhibitor cocktail (Biotool), and 10 μM MG132 for standard SDS-PAGE or Phos-tag SDS-PAGE. Immunoblot analysis was performed as previously described using Blocking One (Nacalai Tesque) or Blocking One-P (Nacalai Tesque) and WesternSure ECL Substrate (Li-Cor Biosciences)46 (link). Protein was visualized by Ez-Capture II (ATTO Corp) and the band intensities were quantified using Image Studio software (Li-Cor Biosciences). The sources of antibodies were as follows: Phospho-Ser51-eIF2α (D9G8) (Cell Signaling Technology); eIF2α (D7D3) (Cell Signaling Technology); anti-PERK (C33E10) (Cell Signaling Technology); anti-GCN2 (Cell Signaling Technology); anti-PKR (M-515) (Santa Cruz Biotechnology); anti-FLAG M2 (Sigma Aldrich); anti-HRI (sc-30143) (Santa Cruz Biotechnology) and anti-HRI (ab84980) (Abcam).
3
Antibody Characterization for Desmosomal and Epidermal Proteins
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Mouse monoclonal antibodies used: P124 (anti–Dsg1 extracellular domain; Progen, Heidelberg, Germany); 27B2 (anti–Dsg1 cytodomain; Invitrogen), U100 (anti-Dsc1; Progen), HECD1 (anti– E-cadherin; Takara, Kyoto, Japan) and 4A4 (anti-p63; Santa Cruz Biotechnology, Dallas, TX). Rabbit polyclonal antibodies used: K1, K10, and loricrin (gifts from J. Segre, National Human Genome Research Institute, Bethesda, MD), 1905 (anti-Dsg3; gift from J. Stanley, University of Pennsylvania, Philadelphia, PA), C33E10 (anti-pERK; Cell Signaling Technology, Danvers, MA), anti-Erk1/2 (Promega, Madison, WI), H-137 (anti-p63; Santa Cruz Biotechnology) and GAPDH (glyceraldehyde-3-phosphate dehydrogenase; Sigma-Aldrich). Chicken polyclonal antibody used: Pg (1407; Aves Laboratories, Tigard, OR). Secondary antibodies for immunoblotting were goat anti–mouse, –rabbit, and –chicken peroxidase (Rockland; KPL, Gaithersburg, MD). Secondary antibodies for immunofluorescence microscopy were goat anti– mouse, –rabbit, and –chicken linked to fluorophores of 488 nm and 568 nm (Alexa Fluor; Invitrogen).
4
Antibody Characterization for Desmosomal and Epidermal Proteins
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