Jejunal tissues and cultured cells were lysed, and the extracted proteins were analyzed. The primary antibodies used for Western blot were rabbit anti-Mst1 (1:1000; Cell Signaling), anti-p-Mst (1:1000; Cell Signaling), anti-Lats1 (1:1000; Cell Signaling), anti-p-Lats (Yu et al. 2010 (link)), anti-YAP (1:1000 [Cell Signaling, #4912], 1:1000 [Santa Cruz Biotechnology, H-9 and 63.7], and 1:1000 [Novus, NB110-58358]), anti-p-YAP (S127; 1:1000; Cell Signaling), anti-p-YAP (S381; 1:1000; Cell Signaling) (Kim et al. 2013 (link)), anti-β-catenin (1:2500; Sigma), anti-YAP/TAZ (1:1000; Cell Signaling), anti-Axin2 (1:1000; Cell Signaling), mouse anti-APC (1:200; Calbiochem, Ab-1), anti-Flag (1:5000; Sigma, M2), anti-HA (1:5000; Sigma), anti-Myc (1:2000; Calbiochem), anti-Actin (1:5000; Millipore), and rat anti-Flag (1:2000; BioLegend, L5). Signals were quantified by ImageJ.
Anti p yap
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-p-YAP is a primary antibody that specifically recognizes the phosphorylated form of the Yes-associated protein (YAP), a key transcriptional regulator in the Hippo signaling pathway. This antibody can be used to detect and quantify the levels of phosphorylated YAP in various cellular and experimental settings.
Automatically generated - may contain errors
Lab products found in correlation
Anti yap, by Cell Signaling Technology (19 mentions)
Anti lats1, by Cell Signaling Technology (7 mentions)
Anti gapdh, by Cell Signaling Technology (5 mentions)
Anti p lats, by Cell Signaling Technology (4 mentions)
Anti yap, by Santa Cruz Biotechnology (4 mentions)
Anti lat, by Cell Signaling Technology (3 mentions)
Anti actin, by Merck Group (2 mentions)
Anti flag, by Merck Group (2 mentions)
Anti yap taz, by Cell Signaling Technology (2 mentions)
Anti p mst, by Cell Signaling Technology (2 mentions)
Anti p lats1, by Cell Signaling Technology (2 mentions)
Anti akt, by Cell Signaling Technology (2 mentions)
Anti ctgf, by Cell Signaling Technology (2 mentions)
Anti lamin b, by Santa Cruz Biotechnology (2 mentions)
Anti cyclin d1, by Cell Signaling Technology (2 mentions)
Anti cdk4, by Abcam (2 mentions)
Anti synaptopodin, by Proteintech (2 mentions)
Anti desmin, by Proteintech (2 mentions)
Anti snail2, by Proteintech (2 mentions)
Anti mst1, by Cell Signaling Technology (2 mentions)
30 protocols using anti p yap
1
Western Blot Analysis of Hippo-YAP Pathway
2
Histological and Immunohistochemical Analysis of Mouse Intestinal Samples
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Mouse intestinal samples were collected, fixed overnight in 4% paraformaldehyde in PBS, embedded in paraffin, and sectioned at 5 µm. Sections were stained with hematoxylin–eosin for histological analysis. Immunohistochemistry on serial sections was performed according to the manufacturers’ protocols. Intraperitoneal injection of 30 mg/kg BrdU (Sigma) dissolved in 3 mg/mL 1× PBS was performed 2 h before tissue harvest. The primary antibodies used for immunohistochemistry were rabbit anti-β-catenin (1:500; Sigma), anti-YAP (1:100; Cell Signaling), anti-p-YAP (S127; 1:100; Cell Signaling), anti-YAP/TAZ (1:100; Cell Signaling), anti-Ki67 (1:1000; Novocastra), anti-Lats1 (1:100; Cell Signaling), and mouse anti-BrdU (1:50; Developmental Studies Hybridoma Bank). The signals were developed using the ABC kit purchased from Vector Laboratories according to the manufacturer's suggestions. Cy3-conjugated goat anti-rabbit and Alexa 488-conjugated goat anti-mouse secondary antibodies (Molecular Probes) were used for immunofluorescence.
3
Western Blot Analysis of Protein Expression
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Protein samples were diluted 1:5 with protein loading buffer (6 × Transgen Biotech, Beijing, China) and heated at 95 °C for 5 min. Protein extracts were separated with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), transferred to polyvinylidene fluoride membranes, and blocked with 5% nonfat dry milk in TBST. The membranes were incubated with primary antibodies at 4 °C overnight and with the horseradish peroxidase (HRP)-conjugated secondary antibodies at 37 °C for 1 hour. Protein bands were visualized by enhanced chemiluminescence reagent (Thermo Fisher Scientific, Waltham, MA) and imaged by a FluorChem M gel documentation system (ProteinSimple, San Jose, CA). The primary antibodies (anti-YAP, anti-p-YAP, anti-LATS1, anti-p-LATS1, anti-MYPT1, anti-p-MYPT1, anti-CREB, anti-p-CREB, anti-GAPDH, anti-α-tubulin, anti-histone H3) and secondary antibodies were obtained from Cell Signaling Technology (Danvers, MA, USA). CTGF antibody was obtained from Sigma-Aldrich (St Louis, MO, USA).
4
Western Blot Analysis of Protein Expression
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Total proteins were extracted with RIPA lysis buffer (Beyotime, Shanghai, China), and electrophoresed using 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis and transferred to polyvinylidene fluoride membranes (Bio-Rad, Hercules, CA, USA). Membranes were blocked in Quick Block (Beyotime, Shanghai, China) for 15 min, then incubated with specific primary antibodies (1:1000) at 4°C overnight. Anti-PAX5 (Biorbyt, Cambridge, UK), anti-slug, anti-E-cadherin, anti-vimentin, anti-p-LATS1, anti-LATS1, anti-p-YAP, and anti-YAP (Cell Signaling Technology, USA) were used. Membranes were incubated in secondary antibodies at room temperature for 1 hour. TBST was used to wash unbound antibodies. An enhanced chemiluminescence detection system was used to detect target proteins. GAPDH and β-actin were internal controls.
5
Antibody Sources for Hippo Pathway
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Rabbit polyclonal MST2 phospho-T336 antibody was previously described (Bae et al., 2017 (link)). Rabbit polyclonal SAV1 phospho-T26 was raised against the SAV1 phospho-peptide with the sequence of VKKEpTSPLLC at an on-campus facility. The following antibodies were purchased from the indicated sources: anti-pMST1/2 (T183/180; GTX133948, GeneTex); anti-STRIP1 (A304-644A) and anti-CCM3 (A304-798A, Bethyl Laboratories Inc); anti-Tubulin (ab4074), anti-SIKE1 (ab121860), anti-STK25 (ab157188) and anti-SLMAP (ab56328, Abcam); anti-MYC (Roche); anti-FLAG (F1804, Sigma); anti-HA (sc-805), anti-PP2AA (sc-6112), anti-SAV1 (sc-101205), anti-STRN3 (sc-13562) and anti-YAP (sc-101199, Santa Cruz Biotechnology); anti-MOB4 (A4590, ABclonal); anti-GM130 (12480), anti-MST1 (3682), anti-MST2 (3952), anti-MST3 (3723), anti-MST4 (3822), anti-GAPDH (2118), anti-MOB1 (13730), anti-pMOB1 (T35;8699), anti-LATS1 (9153), anti-pLATS1/2 (HM; 8654, AL;9157), anti-pYAP (4911), anti-PP2AC (2259), anti-rabbit immunoglobulin G (IgG) (H+L) (Dylight 800 or 680 conjugates), and anti-mouse IgG (H+L) (Dylight 800 or 680 conjugates, Cell Signaling). Alexa Fluor 647 Phalloidin (A22287) was purchased from Thermo Scientific. Phos-tag conjugated acrylamide was purchased from Wako chemicals. Latrunculin B (ab144291) and cytochalasin D (sc-201442) were purchased from Abcam and Santa Cruz Biotechnology, respectively.
6
Investigating NF-κB Signaling Pathway
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PB2 (purity > 90%), DEX (used as a positive control), and LPS derived from bacteria (serotype: 0111:B4, L5293) were purchased from Sigma (St. Louis, MO, USA). Anti-NF-κB, anti-p-NF-κB, anti-Akt, anti-p-Akt, anti-PI3K, anti-p-PI3K, anti-YAP, anti-p-YAP, anti-ROCK, anti-Rhoc, and anti-β-actin antibodies were obtained from Cell Signaling Technology (Danvers, MA, USA). Anti-TLR4 and anti-p-LATS1/2 (phospho-Ser909/872) antibodies were obtained from Santa Cruz Biotechnology (Dallas, TX, USA) and MyBioSource (San Diego, CA, USA), respectively.
7
Western Blot Analysis of Cell Signaling
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The whole-cell extract was obtained in RIPA buffer in the presence of standard protease and phosphatase inhibitors. The protein content was determined with a protein assay reagent (Bio-Rad Laboratories Inc., Hercules, CA, USA), using bovine serum albumin as a standard. Equal protein content of total cell lysates was resolved on polyacrylamide gel (Bolt 4–12% Bis-Tris Plus Invitrogen, Carlsbad, CA, USA), electro-transferred to PVDF membranes (iBlot Invitrogen, Carlsbad, CA, USA) and incubated with specific primary antibodies: anti-YAP (Santa Cruz Biotechnology, Inc., Dallas, TX, USA; sc #101199; dil.1:1000); anti-p-YAP (Cell Signaling Technology, Inc., Beverly, MA, USA; #13008; dil.1:1000); anti-β Actin (MP Biomedicals, Santa Ana, CA, USA; # 69100; dil.1:10000); anti-Rac1, anti-RhoA and anti-Cdc42 (all Cell Biolabs, San Diego, CA, USA; STA-405; dil.1:500); anti-Zeb1 (Abcam, Cambridge, MA, USA; #180905; dil.1:2000); anti-Snail1 (Santa Cruz Biotechnology, Inc., Dallas, TX, USA; sc-271977; dil.1:1000); anti-GSN (Sigma-Aldrich, St Louis, MO, USA; G-4896, dil.1:1000). Membranes were developed using ECL detection reagents (GE Healthcare Life Sciences, Uppsala, Sweden) on a UVITEC imaging system (UVITEC, Cambridge, UK) or ChemiDocMP system (Bio-Rad Laboratories Inc.). Quantitative analysis of western blots was performed using ImageJ (NIH) software [23 (link)].
8
Immunofluorescence Analysis of Carotid Arteries
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The frozen sections of carotid arteries were incubated at 60 °C for 2 min, and the carotid arteries or aortic arches were fixed with 4% paraformaldehyde for 30 min. Then the samples were washed with PBS and permeabilized/blocked with 0.1% Triton-X100 (in 5% BSA). Subsequently, the samples were added with the primary antibody in a wet box at 4 °C overnight. Primary antibodies included anti-G3BP2 (ab86135), anti-CD31 (AF3628), anti-YAP (Cell Signaling Technology, #14074), anti-pYAP (Cell Signaling Technology, #13008), anti-VE-cadherin (sc-9989) and anti-integrin β3 (sc-365679). After the samples were washed with PBST (PBS with 0.1% Tween-20) 5 times, the samples were incubated with secondary antibodies for 1 h and then with DAPI for 10 min at room temperature (protected from light). The fluorescent signal of sections was detected by SP8 confocal microscopy (Lecia, Germany).
9
Quantitative Analysis of Cell Signaling
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Total cell lysates were extracted. Western blotting was performed using total cell lysates. The primary antibodies used in western blot were anti-SAV1 (HPA0018085, Sigma), anti-YAP (#12395, Cell Signaling Technology), anti-KDM2B (SAB2702002, Sigma), anti-pYAP (#13008, Ser127, Cell Signaling Technology), anti-H3K27me3 (#9733, Cell Signaling Technology), and anti-CTGF (#86641, Cell Signaling Technology). Anti-actin (rabbit; Santa Cruz Biotechnology) was used as equal protein-sample loading. Anti-mouse IgG, anti-goat IgG, or anti-rabbit IgG (Santa Cruz Biotechnology) were used as secondary antibodies. Quantity One analysis software was used to quantify the bands (version 4.6; Bio-Rad).
10
Immunoblotting Protein Analysis in Cells
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Immunobloting analysis of proteins in cell lysates and nuclear fractions was performed as previously described [56 (link), 57 (link)]. Primary antibodies used were as follows: anti-YAP (#4912), anti-p-YAP (#4911), anti-Myc-Tag (#2278), anti-Mst (#3682), anti-p-Mst (#3681), anti-LATS (#3477) and anti-p-LATS (#8654) were purchased from Cell Signaling Technology (Beverly, MA); anti-HIF-1α (#610959) was purchased from BD; anti-β-Actin (#Sc-47778), anti-Lamin B (#Sc-6216), anti-BNIP3 (#Sc-56167), and anti-HMGCR (#27578) were purchased from Santa Cruz (Santa Cruz, CA).
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