RIPA (GBCBIO, G3424) was used to lyse cells and extract total protein to prepare protein samples. The protein lysates were separated by conventional electrophoresis, transferred onto the membrane, and washed with PBS, followed by blocking with 1% BSA for 2 h. Then, primary antibodies Box (ab32503, Abcam, Cambridge, UK), Bcl-2 (ab32124, Abcam, Cambridge, UK), Caspase-1 (ab32503, Abcam, Cambridge, UK), Cleaved Caspase-3 (ab32503, Abcam, Cambridge, UK), Caspase-8 (ab207802, Abcam, Cambridge, UK), Caspase-9 (ab2302, Abcam, Cambridge, UK), Caspase-11 (ab25901, Abcam, Cambridge, UK), Cyclin D1 (ab202068, Abcam, Cambridge, UK), ERK 1/2 (ab180673, Abcam, Cambridge, UK), p-ERK 1/2 (ab16663, Abcam, Cambridge, UK), WNT (ab17942, Abcam, Cambridge, UK), β-catenin (ab223500, Abcam, Cambridge, UK), GSDMD (ab15251, Abcam, Cambridge, UK) and β-actin (ab32572, Abcam, Cambridge, UK) (abcam, ab32503, ab32124, ab207802, ab2302, ab25901, ab202068, ab180673, ab16663, ab17942, ab223500, ab15251, ab32572, ab219800, ab8227) each at 1:1000 dilution were incubated with the membrane overnight at 4°C. The next day, HRP (ab32572, Abcam, Cambridge, UK) and labeled IgG secondary antibody (ab150077, Abcam, Cambridge, UK) each at 1:5,000 were further incubated with the membrane at room temperature for 1 h. ECL Luminescence Kit (Thermo, 32209) was used to develop the signal in a dark room.
Ab17942
Manufactured by Abcam
Sourced in United States, United Kingdom, China
Ab17942 is a recombinant monoclonal antibody targeting the Epidermal Growth Factor Receptor (EGFR) protein. This product is designed for use in various research applications.
Automatically generated - may contain errors
Lab products found in correlation
Ab50011, by Abcam (16 mentions)
Ab205718, by Abcam (12 mentions)
Ab214362, by Abcam (12 mentions)
Ab223500, by Abcam (11 mentions)
Ab8245, by Abcam (9 mentions)
Pvdf membrane, by Merck Group (9 mentions)
Ab181602, by Abcam (8 mentions)
Ab124956, by Abcam (8 mentions)
Ab31828, by Abcam (8 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (8 mentions)
Bca protein assay, by Thermo Fisher Scientific (8 mentions)
Ab201015, by Abcam (6 mentions)
Ab179461, by Abcam (6 mentions)
Ab8227, by Abcam (6 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (6 mentions)
Ab47363, by Abcam (5 mentions)
Ab178876, by Abcam (5 mentions)
Ab8805, by Abcam (5 mentions)
Ab16502, by Abcam (5 mentions)
Ab32503, by Abcam (5 mentions)
92 protocols using ab17942
1
Western Blot Analysis of Apoptosis Markers
2
Protein Expression Analysis in RAW264.7 Cells
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Total protein was extracted from tissues or RAW264.7 cells using Radio Immunoprecipitation Assay (RIPA) Lysis Buffer (R0010; Solarbio). After SDS‐PAGE separation and membrane transfer, immunoblots were obtained by using the following primary rabbit antibodies: GAPDH (#5174, 1:1000, Rabbit, Cell Signaling Technology), TLR4 (ab13556, 1:500; Abcam Inc.), phosphorylated‐ERK1/2 (p‐ERK1/2) (ab17942, 1:1000; Abcam), ERK1/2 (ab17942, 1:1000; Abcam), KLF4 (ab129473, 1:1000; Abcam) and ITGA2B (ab134131, 1:2000; Abcam). The immunoblots were visualized with secondary goat anti‐rabbit antibody to immunoglobulin G (ab150077, 1:1000; Abcam) and enhanced chemiluminescence, and analysed by Image J software. The relative expression of protein was expressed as the ratio of grey value of the target band to that of internal reference (GAPDH).
3
Comprehensive Antibody Panel for EMT Analysis
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Antibodies against HMGB3 (27465-1-AP), ETS-1 (12118-1-AP), MEK1/2 (11049-1-AP), CCND1 (60186-1-Ig), c-Myc (10828-1-AP), SOX2 (11064-1-AP), and ALDH1A1 (15910-1-AP) were purchased from Proteintech (Wuhan, China). Antibodies for p-MEK1/2 (9154), p-ERK1/2 (4370), and an Epithelial-Mesenchymal Transition Antibody Sampler Kit (9782) were obtained from Cell Signaling Technology (Danvers, MA, USA). Antibody against ERK1/2 (ab17942) was obtained from Abcam (Cambridge, UK). Antibody against β-actin (A5441) was purchased from Sigma-Aldrich (St. Louis, MO, USA). AZD6244 (S1008) and PD0325901 (S1036) were acquired from Selleck Chemicals (Houston, TX, USA).
4
Angiogenesis Signaling Pathway Protocol
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All cell culture serum and media were purchased from Thermo Fisher Scientific, while cell culture supplements were purchased from Sigma. Rat anti-CD31 (553369) was purchased from Pharmingen while antibodies against alpha SMA (ab7817), phosphor-ErK (ab50011) and ErK1/2 (ab17942) were from Abcam. The antibodies against phospho-Akt (sc-7985R), Akt1 (sc-1619) and GAPDH (sc-25778) were from Santa CruZ Biotech. The antibody against VEGF (SAB1402390) was from Sigma. All secondary antibodies were from Dakocytomation. All microRNA reagents were purchased from Thermo Fisher Scientific. PDGF, insulin, holo-transferrin, SU5416, LY294002, DMSO, DAPI and Giemsa were purchased from Sigma.
5
Protein Expression Analysis in Cells
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The content of PC1, SM22, ACTA2, CNN1, p38, ERK, JNK, PI3K and Myc, CyclinD1, and the phosphorylation of p38, ERK, JNK and PI3K were determined by Western blot. Preparation of whole cell lysates and tissue homogenates, and the immunoblotting assay were performed according to previously described procedures.18 (link) The primary antibodies were obtained from the following sources: anti-Polycystin-1 (ABT128; Millipore); anti-SM22 alpha (ab14106; Abcam, Cambridge, UK); anti-alpha smooth muscle Actin (ab5694; Abcam); anti-Calponin (ab46794; Abcam); anti-alpha Tubulin (ab52866; Abcam); anti-p38 (phospho Y182) (ab47363; Abcam); Anti-p38 (ab7952; Abcam); Anti-ERK1 (pT202/pY204) + ERK2 (pT185/pY187) (ab50011; Abcam); Anti-ERK1 + ERK2(ab17942; Abcam); Anti-JNK1+JNK2 (phospho T183 + Y185) antibody (ab4821); Anti-JNK1+JNK2 (ab37228; Abcam); Anti-PI3K p85 (phospho Y607); Anti-PI3K p85 (ab189403; Abcam); anti-myc (ab32072; Abcam) and anti-Cyclin D1 (ab134175; Abcam).
6
Ovarian Cancer Cell Protein Analysis
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Ovarian cancer cells were disrupted using cell lysis buffer (Promega) on ice for 30 min and centrifuged at 12000 rpm for 30 min. The supernatant was transferred into the new centrifuge tube, and the concentration of protein samples was detected using the BCA-200 Protein Assay kit (Pierce, Rockford, IL, USA). Protein samples were separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) gel and transferred to the polyvinylidene fluoride (PVDF) membrane (Millipore). The non-specific sites in the membrane were blocked using 5% skim milk for 1 h, followed by incubation with primary antibodies and horseradish peroxidase (HRP)-labeled secondary antibody. The blots were visualized using the enhanced chemiluminescent (ECL) system (Beyotime, Shanghai, China). The primary antibodies, including phosphorylated extracellular regulated MAP kinase (p-ERK; ab214036), ERK (ab17942), p-MAP kinse-ERK kinase (p-MEK; ab96379), MEK (ab178876) and GAPDH (ab181602) were purchased from Abcam (Cambridge, MA, USA).
7
Western Blot Analysis of Hippo Pathway
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Anti-VEGF antibody, Abcam (Cambridge, UK), ab1316, mouse monoclonal, western blot (WB) dilution: 1:250; anti-extracellular signal-regulated kinas (Erk1) (pT202/pY204) + Erk2 (pT185/pY187) antibody, Abcam, ab4819, rabbit polyclonal, western blot (WB) dilution: 1:1000; anti-Erk1/2 antibody, Abcam, ab17942, rabbit polyclonal, western blot (WB) dilution: 1:1000; anti-Lats antibody, Santa Cruz (Dallas, Texas, U.S.A.), sc-9388, goat polyclonal, western blot (WB) dilution: 1:200; anti-TAZ antibody, Santa Cruz, sc-48,805, rabbit polyclonal, western blot (WB) dilution: 1:500; anti-TEAD antibody, Santa Cruz, sc-134,070, rabbit polyclonal, western blot (WB) dilution: 1:1000; anti-β-actin antibody, Abcam, ab8226, mouse monoclonal, western blot (WB) dilution: 1:5000; P-MST, CST (Boston, Massachusetts, USA), #3681, rabbit polyclonal, immunohistochemistry dilution: 1:250; P-YAP, CST, # 4911, rabbit polyclonal, immunohistochemistry dilution: 1:250.
8
Comprehensive Antibody Panel for Cell Signaling
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Mouse anti‐AKAP95 (sc‐100 643), anti‐Cx43 (sc‐13 558), anti‐cyclin D2 (sc‐53 637), anti‐cyclin D3 (sc‐6283), and anti‐Cdk2 (sc‐70 829) antibodies were purchased from Santa Cruz Biotechnology (TX, USA). Rabbit anti‐cyclin D1 (ab134175), anti‐PKB (ab8805), anti‐PKC (ab32376), anti‐ERK1/2 (ab17942), anti‐ERK5 (ab40809), anti‐AKAP95 (ab140628), anti‐cyclin E1 (ab33911), and anti‐cyclin E2 (ab40890) antibodies were purchased from Abcam (Cambridge, UK). Rabbit anti‐cyclin D1‐T286 (3300S) antibody was purchased from Cell Signaling Technology (MA, USA). Mouse anti‐GAPDH (D190090‐O100) was purchased from BBI Life Sciences (Shanghai, China).
9
Western Blot Analysis of Cell Signaling Pathways
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Total protein
extracts were generated using lysis buffer (50 mM Tris-HCl, pH 7.4,
150 mM NaCl, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 1
mM EDTA) supplemented with PhosSTOP and Complete Phosphatase/Protease
Inhibitor Cocktails (Roche Diagnostics GmbH, Mannheim, Germany). Protein
extracts (20–25 μg per sample) were loaded onto sodium
dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels
and transferred electrophoretically to poly(vinylidene fluoride) (PVDF)
membranes. The following primary antibodies were used at a dilution
of 1:1000: ERK1/2 (p44/42) (ab17942, Abcam), p-ERK1/2 (p-p44/42) (Thr202/Tyr204)
(9101, Cell Signaling), MNK1 (2195, Cell Signaling), p-MNK1 (Thr197/202)
(2111, Cell Signaling), eIF4E (9742, Cell Signaling), p-eIF4E (p-Ser
209) (9741, Cell Signaling or NBP2-66802, Novus Biologicals), p38alpha
(9218, Cell Signaling), p38beta (2339, Cell Signaling), p-p38 (4511,
Cell Signaling), p-ATF2 (27934, Cell Signaling), Hsp27 (2402, Cell
Signaling), p-Hsp27 (S82) (2401, Cell Signaling), and p-p90 RSK (Thr573)
(9346, Cell Signaling). The primary HRP-conjugated antibody anti-β-actin
(Calbiochem) was used at a dilution of 1:20 000. Anti-mouse
and anti-rabbit HRP secondary antibodies were from Pierce and used
at a dilution of 1:10 000. Immunodetection of proteins was
performed using ECL Western Blotting Detection Reagents (GE Healthcare,
Buckinghamshire, U.K.).
extracts were generated using lysis buffer (50 mM Tris-HCl, pH 7.4,
150 mM NaCl, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 1
mM EDTA) supplemented with PhosSTOP and Complete Phosphatase/Protease
Inhibitor Cocktails (Roche Diagnostics GmbH, Mannheim, Germany). Protein
extracts (20–25 μg per sample) were loaded onto sodium
dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels
and transferred electrophoretically to poly(vinylidene fluoride) (PVDF)
membranes. The following primary antibodies were used at a dilution
of 1:1000: ERK1/2 (p44/42) (ab17942, Abcam), p-ERK1/2 (p-p44/42) (Thr202/Tyr204)
(9101, Cell Signaling), MNK1 (2195, Cell Signaling), p-MNK1 (Thr197/202)
(2111, Cell Signaling), eIF4E (9742, Cell Signaling), p-eIF4E (p-Ser
209) (9741, Cell Signaling or NBP2-66802, Novus Biologicals), p38alpha
(9218, Cell Signaling), p38beta (2339, Cell Signaling), p-p38 (4511,
Cell Signaling), p-ATF2 (27934, Cell Signaling), Hsp27 (2402, Cell
Signaling), p-Hsp27 (S82) (2401, Cell Signaling), and p-p90 RSK (Thr573)
(9346, Cell Signaling). The primary HRP-conjugated antibody anti-β-actin
(Calbiochem) was used at a dilution of 1:20 000. Anti-mouse
and anti-rabbit HRP secondary antibodies were from Pierce and used
at a dilution of 1:10 000. Immunodetection of proteins was
performed using ECL Western Blotting Detection Reagents (GE Healthcare,
Buckinghamshire, U.K.).
10
Immunohistochemical Analysis of MAPK Signaling
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The sections of colon tissue were deparaffinized by xylene, hydrated with gradient ethanol, incubated with 0.1% Tritonx-100 for 30 min, and rinsed 3 times with PBS for 5 min each. The sections were blocked by 5% BSA and 10% sheep serum successively for 30 min. The sections were incubated separately with primary antibody ERK1/2 (ab17942, Abcam), p-ERK (ab192591, Abcam), p38 (ab31828, Abcam), p-p38 (ab47363, Abcam), JNK (ab208035, Abcam), and p-JNK (ab124956, Abcam) in a wet box at 4°C overnight, washed 3 times with PBS for 5 min per time and second antibody at 37°C for 40 minutes, and washed with PBS 3 times for 5 minutes per time. Horseradish peroxidase-labeled streptavidin protein was added dropwise at 37°C for 40 minutes. The sections were rinsed 3 times with PBS for 5 minutes each time, developed by DAB, microscopically observed, and photographed.
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