Following cell lysis by ice-cold radioimmunoprecipitation assay buffer, protein concentrations were determined. Equal quantities (30 µg) of protein were separated using 10% SDS-polyacrylamide gels, transferred to a polyvinylidene difluoride (PVDF) membrane and blocked with 5% bovine serum albumin (BSA). PVDF membranes were incubated overnight at 4°C with the following antibodies: CIP2A (dilution 1:500; cat. no. sc-80662; Santa Cruz Biotechnology, Inc., Dallas, CA, USA), AKT (dilution 1:1,000; cat. no. 9272S; Cell Signaling Technology, Inc., Danvers, MA, USA), phosphorylated (p)-AKT (dilution 1:1,000; cat. no. 4060S; Cell Signaling Technology), B cell lymphoma-2 (Bcl-2; dilution 1:1,000; cat. no. sc-492; Santa Cruz Biotechnology, Inc.), caspase-3 (dilution 1:1,000; cat. no. 9662S; Cell Signaling Technology), poly(ADP-Ribose) polymerase (PARP; dilution 1:1,000; cat. no. 5625T; Cell Signaling Technology), γ-H2AX (dilution 1:1,000; cat. no. ab2893; Abcam, Cambridge, UK) and β-actin (dilution 1:1,000; cat. no. sc-47778; Santa Cruz Biotechnology, Inc.). After being washed, the membranes were then incubated with horseradish peroxidase-conjugated secondary antibodies (anti-mouse IgG, dilution 1:1,000; cat. no. 7076S; Cell Signaling Technology; anti-rabbit IgG, dilution 1:1,000; cat. no. 7074S; Cell Signaling Technology) followed by enhanced chemiluminescence detection.
B cell lymphoma 2 bcl 2
Manufactured by Cell Signaling Technology
Sourced in United States, China
B-cell lymphoma-2 (Bcl-2) is a protein that plays a crucial role in the regulation of apoptosis, or programmed cell death. It functions as an anti-apoptotic protein, promoting cell survival by inhibiting the activity of pro-apoptotic proteins. Bcl-2 is an important target in cancer research and drug development, as its dysregulation is commonly observed in various types of cancer.
Automatically generated - may contain errors
Lab products found in correlation
Bcl2 associated x (bax), by Cell Signaling Technology (7 mentions)
Bcl 2 associated x protein bax, by Cell Signaling Technology (5 mentions)
Protease inhibitor cocktail, by Thermo Fisher Scientific (4 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (4 mentions)
Ripa lysis buffer, by Thermo Fisher Scientific (4 mentions)
Cleaved caspase 3, by Cell Signaling Technology (3 mentions)
Caspase 3, by Cell Signaling Technology (3 mentions)
C myc, by Cell Signaling Technology (3 mentions)
β actin, by Cell Signaling Technology (3 mentions)
Nlrp3, by Novus Biologicals (2 mentions)
Ecl system, by Thermo Fisher Scientific (2 mentions)
Caspase 8 cas8, by Cell Signaling Technology (2 mentions)
Il 1β, by Abcam (2 mentions)
Caspase 1, by Santa Cruz Biotechnology (2 mentions)
Gapdh, by Merck Group (2 mentions)
β actin, by Merck Group (2 mentions)
P mtor, by Cell Signaling Technology (2 mentions)
Mtt reagent, by Merck Group (2 mentions)
Gapdh, by Proteintech (2 mentions)
Bcl 2 associated x protein bax, by Abcam (2 mentions)
25 protocols using b cell lymphoma 2 bcl 2
1
Western Blot Analysis of Apoptosis Markers
2
Western Blot Analysis of Rat Hippocampus
3
Western Blot Analysis of Oxidative Stress Markers
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Kidney tissues of sham operated, saline treated and fetal kidney cells treated rats were homogenised in RIPA buffer containing 1 mmol/L phenylmethanesulphonyl fluoride (PMSF) and 1% protease inhibitor cocktail (Sigma-Aldrich, MO, USA). 40 mg proteins were loaded and separated on 10% sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE). After electrophoresis, separated proteins were transferred to polyvinylidene difluoride (PVDF) membranes. Membranes were blocked with 5% BSA for 1 hour at room temp and incubated overnight at 4°C with appropriate dilutions of primary antibodies: nuclear factor-κB (NFκB), intercellular adhesion molecule-1 (ICAM-1), heme oxygenase-1 (HO-1), NAD(P)H quinone oxidoreductase 1 (NQO-1), cytochrome c, caspase 3 (all from Abcam, MA, USA), B-cell lymphoma 2-associated X protein (Bax) and B-cell lymphoma 2 (Bcl2) (both from Cell Signaling Technology, MA, USA). β-actin antibody (Abcam, MA, USA) was used as a loading control. After incubation, PVDF membranes were washed three times with TBS and incubated with corresponding horse raddish peroxidase-conjugated secondary antibody for 1 hour at room temperature. All proteins were detected using super signal west pico chemiluminescent substrate (Thermo scientific, IL, USA) and quantified by densitometry using the Quantity One software (Bio-Rad, CA, USA).
4
Quantification of Protein Levels in Neuronal Cells
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Protein levels in the extracts of SH-SY5Y cells or the hippocampal tissues were analyzed using western blot as previously described [5] (link) with primary antibodies against Thr205-phosphorylated tau-, Tyr307-phosphorylated protein phosphatase 2 A (PP2A)-, total PP2A (Santa Cruz Biotechnology, Santa Cruz, CA, USA) and Thr181-phosphorylated tau-, total tau-, Thr202/Tyr204-phosphorylated extracellular signal-regulated kinases 1/2 (ERK1/2)-, Thr180/Tyr182-phosphorylated p38 mitogen activated protein kinase (p38 MAPK)-, Thr183/Tyr185-phosphorylated c-Jun N-terminal kinase (JNK)-, β-actin-, total ERK1/2-, total p38-, total JNK-, hypoxia-inducible factor-1α (HIF-1α)-, B-cell lymphoma-2 (Bcl-2)-, Bcl2-associated X (Bax)-, cleaved caspase-3-specific antibodies (Cell Signaling Technology, Beverly, MA, USA). Immunoreactive bands were detected with the appropriate horseradish peroxidase-conjugated secondary antibodies (Cell Signaling Technology) and immunological complexes were visualized by enhanced chemiluminescence reagents (Millipore, Billerica, MA, USA). Western blot signals were analyzed by ImageJ software.
5
Comprehensive Antibody Panel for Western Blot and IHC
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For western blotting and immunohistochemistry analyses, the following antibodies were purchased from Abcam (Cambridge, UK): anti-H2A.Z (cat no. ab150402; dilution 1:1,000 for western blot analysis and dilution 1:300 for immunohistochemistry), anti-p21/WAF1/Cip1 (cat. no. ab109520), anti-p27/Kip1 (cat. no. ab32034), anti-Skp2 (cat. no. ab124799), anti-cyclinA (cat. no. ab181591) and matrix metalloproteinase (MMP)2 (cat. no. ab37150). The following antibodies were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA): anti-cyclin-dependent kinase (CDK) 4 (cat. no. 12790), anti-CDK6 (cat. no. 3136) and anti-CDK2 (cat. no. 2546). Ki67 (cat. no. ab15580; Abcam) was used for the immunohistochemical analysis of mouse tumor tissue. The following antibodies were purchased from Epitomics (Burlingame, CA, USA): anti-β-actin (cat. no. 1854-1), and anti-GAPDH (cat. no. 5632-1). A kit with antibodies targeting EMT-associated proteins (E-cadherin, N-cadherin, Slug, Snail, and Vimentin) was obtained from Cell Signaling Technology, Inc. (cat, no. 9782S). Antibodies, recognizing total (#9665) and cleaved caspase-3 (#9664), caspase-9 (#9508), Bcl-2 homologous antagonist/killer (Bak) (#12105) and B-cell lymphoma 2 (Bcl-2) (#15071) were obtained from Cell Signaling Technology, Inc.
6
Resveratrol and Chloroquine Modulate Autophagy
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Resveratrol and chloroquine were purchased from Sigma-Aldrich (Merck Millipore, Darmstadt, Germany). The H9c2 cells were obtained from the Type Culture Collection of the Chinese Academy of Sciences (Shanghai, China). Lactate dehydrogenase (LDH) activity assay and MTT kits were purchased from Beyotime Institute of Biotechnology (Shanghai, China). Primary antibodies against Cx43 (cat no. 3512), AMP-protein activated kinase (AMPK; cat no. 2532) and phosphorylated (p)-AMPK (cat no. 2531S), mammalian target of rapamycin (mTOR; cat no. 2972), p-mTOR (cat no. 2971), Beclin-1 (cat no. 3738), p62 (cat no. 5114), LC3-I/II (cat no. 2775), B-cell lymphoma-2 (Bcl-2; cat no. 2870) and Bcl-2-associated X protein (Bax; cat no. 2772) were obtained from Cell Signaling Technology, Inc. (Danvers, MA, USA).
7
Quantification of Apoptosis Markers in Cells
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XHP was purchased from Tong Ren Tang Technologies Co., Ltd. (Beijing, China). Dimethyl sulphoxide and MTT reagent were purchased from Sigma-Aldrich; Merck KGaA (Darmstadt, Germany). Human insulin, epidermal growth factor, cholera toxin and hydrocortisone were purchased from Gibco; Thermo Fisher Scientific, Inc. (Waltham, MA, USA). The human caspase-3 monoclonal antibody (cat. no. ab32351; 1:3,000 dilution) was purchased from Abcam (Cambridge, UK). The human cyclin A (cat. no. 4656; 1:5,000 dilution), p21Cip1 (cat. no. 2947; 1:5,000 dilution), caspase-8 (cat. no. 9746; 1:4,000 dilution), Bcl-2-associated X protein (Bax; cat. no. 5023; 1:5,000 dilution) and B-cell lymphoma 2 (Bcl-2; cat. no. 2870; 1:5,000 dilution) monoclonal antibodies were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). The β-actin rabbit monoclonal antibody (cat. no. TDY051; 1:5,000 dilution) was purchased from Beijing TDY Biotech Co., Ltd. (Beijing, China). The horseradish peroxidase-conjugated goat anti-rabbit IgG secondary antibodies (cat. no. ZB-2306; 1:5,000 dilution) and goat anti-mouse IgG secondary antibodies (cat. no. ZF-0312; 1:5,000 dilution) were purchased from OriGene Technologies, Inc. (Beijing, China).
8
Protein Expression Analysis by Western Blotting
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Western blotting was performed as previously described (Wang et al., 2015 (link)). Briefly, cell protein extracts (15–20 μg) were separated by 10–15% SDS-PAGE and transferred to PVDF membranes. Proteins were detected with the following antibodies: SIRT3 (1:1000, Cell Signaling Technology, Boston, MA, USA), MnSOD (1:1000, Proteintech, Chicago, IL, USA), cyclophilin D (CypD; 1:1000, Abcam, Cambridge, UK), cytochrome C (Cyt C; 1:1000, Cell Signaling Technology), cleaved caspase-3 (1:1000, Cell Signaling Technology), Bax (1:1000, Cell Signaling Technology, Boston, MA, USA), B-cell lymphoma 2 (Bcl-2; 1:1000, Cell Signaling Technology, Boston, MA, USA), COX-IV (1:1000, Abcam, Cambridge, MA, USA) and GAPDH (1:2000, Proteintech, Chicago, IL, USA).
9
Protein Extraction and Western Blot Analysis
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The cells were lysed in the radioimmunoprecipitation assay (RIPA) lysate of a total protein extraction kit (Bioswamp, China), and the protease inhibitor phenylmethylsulfonyl fluoride (PMSF) (Servicebio, China) and a cocktail (Servicebio, China) were added to obtain protein extracts. Then, 40 μg of protein from each group was added to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels and transferred to polyvinylidene fluoride (PVDF) membranes. The membranes were incubated with primary antibodies against GSDME (#AB215191, Abcam), caspase-9 (#9502, Cell Signaling Technology), caspase-3 (#9662, Cell Signaling Technology), caspase-7 (#12827, Cell Signaling Technology), B-cell lymphoma-2 (Bcl-2) (#4223, Cell Signaling Technology), Bcl2-associated X (Bax) (#2774, Cell Signaling Technology), and GAPDH (#60004, Proteintech, China) overnight at 4°C. Then, the membranes were washed with Tris-buffered saline and tween 20 (TBST) and incubated with horseradish peroxidase- (HRP-) labelled secondary antibodies (#7074, Cell Signaling Technology). The protein bands were visualized with Image Lab software (Bio-Rad, USA).
10
Protein Expression Analysis of Key Metabolic Regulators
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The harvested cells were lysed with radioimmunoprecipitation assay buffer (Beyotime, P0013B) for the extraction of total protein. Protein concentration was quantified with enhanced bicinchoninic acid protein assay kit (Beyotime, P0010). Then, lysate protein was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis and electrophoretically transferred to polyvinylidene difluoride membranes (Millipore, ISEQ00010). The membranes were sequentially blocked and incubated overnight with the following primary antibodies: HK2 (Cell Signaling Technology, 2106), platelet-type phosphofructokinase (Cell Signaling Technology, 8164), phosphoglycerate kinase 1 (Abcam, ab38007), PKM2 (Cell Signaling Technology, 4053), LDHA (Cell Signaling Technology, 3582), c-Myc (Cell Signaling Technology, 13987), B-cell lymphoma-2 (Bcl-2) (Cell Signaling Technology, 4223), Bcl-XL (Proteintech, 26967-1-AP), Bad (Cell Signaling Technology, 9239), Bax (Cell Signaling Technology, 9292) and β-actin (Sungene Biotech, KM9006). The protein bands were acquired as an electronic images format using a ChemiDocTM XRS+ System (Bio-Rad) with immobilon western chemiluminescent horseradish peroxidase substrate (Millipore, WBKLS0100) and quantified the intensities by Image Lab software.
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