Expression levels of apoptosis-related proteins including Bcl-2 (Transduction, Bluegrass-Lexington, KY), Bax (Chemicon, Temecula, CA), caspase 3 (CPP32, Upstate Biotechnology, Lake Placid, NY), PARP (Cell Signaling Technology, Inc., Danvers, MA), and β-actin (Sigma, Saint Louis, MI) were analyzed using Western blotting in the duodenal epithelium from the ulcers of rats in the different groups [24] (link).
Bcl2 associated x (bax)
Manufactured by Merck Group
Sourced in United States, Germany
Bax is a laboratory equipment product manufactured by Merck Group. It is designed to perform essential functions in research and analytical settings. The core function of Bax is to provide reliable and accurate measurements, but a detailed description while maintaining an unbiased and factual approach is not available.
Automatically generated - may contain errors
Lab products found in correlation
Bcl 2, by Merck Group (32 mentions)
β actin, by Merck Group (20 mentions)
Gapdh, by Merck Group (11 mentions)
Caspase 3, by Merck Group (10 mentions)
Cleaved caspase 3, by Merck Group (10 mentions)
Bcl xl, by Merck Group (6 mentions)
Caspase 9, by Merck Group (4 mentions)
Bcl 2, by Santa Cruz Biotechnology (4 mentions)
Cleaved caspase 3, by Cell Signaling Technology (4 mentions)
Mcl 1, by Cell Signaling Technology (4 mentions)
Bcl 2, by Cell Signaling Technology (4 mentions)
Nf κb, by Merck Group (4 mentions)
Bcl xl, by Cell Signaling Technology (4 mentions)
Pvdf membrane, by Merck Group (3 mentions)
Trypsin, by Merck Group (3 mentions)
Erk1 2, by Merck Group (3 mentions)
Beclin 1, by Merck Group (2 mentions)
Gapdh, by Santa Cruz Biotechnology (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Cyclin d1, by Merck Group (2 mentions)
62 protocols using bcl2 associated x (bax)
1
Apoptosis Protein Expression in Duodenal Ulcers
2
Phillyrin Modulation of Apoptotic Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blot was performed as previously reported [25 (link)]. In brief, primary neuron cells were incubated with 20, 40, and 80 μM phillyrin for 2 h followed by 30 min treatment with 100 μM H2O2. Total proteins were extracted from these cells and transferred onto cellulose acetate membranes after separated by SDS-PAGE. After blocking with PBS solution for 1 h, the membranes were incubated with Bax (1:1000, Chemicon, Temecula, CA, USA), Bcl-2 (1:1000), and procaspase-3 (1:1000) primary antibodies from Chemicon, Temecula, CA, USA; pAkt-1 (1:1000), Akt-1 (1:1000), mTOR (1:1000), and LC3-II (1:1000) from Cell Signaling Technology, Danvers, MA, USA; and beclin-1 (1:1000) and β-actin (1:1000) primary antibodies from Sigma-Aldrich, St. Louis, MO, USA, at 4°C for overnight. Then, secondary antibody was added for 1 h at 25°C. ECL chemiluminescence solution was used to visualize protein bands and analyzed using Image J software.
3
Immunoblot Analysis of Apoptosis Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Twenty micrograms of total protein was separated by 12.5% sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to a polyvinylidene fluoride (PVDF) membrane (Millipore, Billerica, MA, USA) for immunoblot analysis using the following primary antibodies: GRP78 (1:1000), GADD153 (1:1000), Bax (1:2000), Bcl-2 (1:1000), cytochrome c (1:500), caspase-9 (1:1000), caspase-3 (1:1000), and GAPDH (1:1000). The primary antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA), except for the Bcl-2 and GAPDH antibodies, which were purchased from Santa Cruz Biotechnology (Dallas, TX, USA). The PVDF membranes were incubated overnight at 4 °C with the primary antibodies, and subsequently incubated for 1 h at room temperature with a horseradish peroxidase (HRP)-conjugated secondary antibody, goat anti-rabbit immunoglobulin G (IgG) or horse anti-mouse IgG (Cell Signaling Technologies, Danvers, MA, USA). The target protein bands were detected using a chemiluminescence image analyzer (CAS-400SM, Davinch-K, Seoul, Korea) and quantified using the ImageJ software (National Institutes of Health, Bethesda, MD, USA). The protein levels were normalized to the GAPDH protein levels.
4
Immunoblotting of Mitochondrial Proteins
5
Western Blot Analysis of THP-1 Macrophages
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
THP-1 derived macrophages were homogenized in 100 μl of protein extraction reagent (Thermo Scientific) and protease inhibitor (Panomics). Protein concentration was determined by Pierce BCA Protein Assay Kit (Thermo Scientific). 20 μg protein was loaded per lane and separated by NuPAGE Novex Bis-Tris 4–12 % mini gel electrophoresis (Invitrogen) in the Novex Xcell-II apparatus for 120 min at 100 V, and transferred to Immbilon-PVDF transfer membranes (Millipore) for immunoblotting. Proteins were visualized by enhanced chemiluminescence according to the manufacturer's instruction. Nonspecific binding was blocked with 5% nonfat milk for 1 h at the room temperature. The antibodies to IKKα, IKKβ, IKKγ, IκBα, p-IκBα and β-actin were purchased from Genetex. The antibodies to phosphorylated-IKKα (phospho Ser176/Ser180), phosphorylated IKKβ (phospho Ser177/Ser181), phosphorylated IKKγ (Ser85) were purchased from Biocompare. The antibody to IκBβ was purchased from Abcam. The antibody to p-IκBβ was purchased from Cell signaling. The antibodies to Bcl-2 and FasL were purchased from BD Biosciences (Pharmingen). The antibodies to p50, p52, Rel A, Rel B and Lamin B were purchased from Santa Cruz. The antibody to p-53 was purchased from Sigma. The antibody to Noxa was purchased from Calbiocam. The antibodies to ABCA1, PUMA, Bax, activated caspase3 and GAPDH were purchased from Millipore.
6
Protein Extraction and Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The cells were collected and lysed using RIPA buffer (Beyotime, Shanghai, China) with a protease inhibitor cocktail (APExBIO, TX, USA) on ice for 20 min. Next, the suspension was centrifuged at 14,000 rpm at 4 °C for 20 min. The supernatant was transferred into new microcentrifuge tubes (Eppendorf, Hamburg, Germany) and the protein concentration was detected using a BCA protein assay kit (Thermo Fisher scientific, CA, USA). Twenty µg of each sample was used for SDS-PAGE. The following primary antibodies were used: E-cadherin (Abcam, Cambridge, MA, USA, ab194982) Vimentin (Abcam, ab193555), N-cadherin (Abcam, ab202030), cyclinD1 (Millipore, Billerica, MA, USA, ABE52), cleaved-casepase3 (Millipore, PC679), bax (Millipore, MAB4601), bcl-2 (Abcam, ab185002), and c-myc (Abcam, ab190026).
7
Protein Expression Analysis of Lung Tissue
8
Whole cell lysate immunoblotting protocol
9
Histopathological and Apoptosis Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
At the moment of sacrificing, the left kidney was taken for the morphological analysis. Renal tissue was prepared as described previously [8] (link), and stained by hematoxiline eosine (H&E) and periodic acid-Schiff (PAS). Intensity and spread of tubular necrosis, number of intra-luminal cast formations, swelling and vacuolization of cells, loss of luminal membrane or brush borders, tubular dilatation, interstitial oedema and separation of cells from tubular basal membrane were semi-quantitatively evaluated as described previously [8] (link). The level of each manifestation was graded with 1 for low, 2 for moderate, 3 for high, and 0 for the lack of manifestation. The sum of these changes represented the histopathological score.
For investigation of Bax and Bcl-2 expression paraffin, sections were treated by microwave for 20 min at 400 W in citrate buffer (pH 6.0) after deparaffinization and dehydration. After antigen retrieval, samples were incubated with Bax (dilution 1∶250, Millipore, Billerica, MA) and Bcl-2 (dilution 1∶200, Millipore, Billerica, MA) antibodies for 1 hour at room temperature. The EnVisionTM staining method (DAKO) was performed, followed by counterstaining with hemalaun (Merck). Negative controls were performed by omitting the first antibody. The slides were evaluated using the light microscope BX53 (Olympus).
For investigation of Bax and Bcl-2 expression paraffin, sections were treated by microwave for 20 min at 400 W in citrate buffer (pH 6.0) after deparaffinization and dehydration. After antigen retrieval, samples were incubated with Bax (dilution 1∶250, Millipore, Billerica, MA) and Bcl-2 (dilution 1∶200, Millipore, Billerica, MA) antibodies for 1 hour at room temperature. The EnVisionTM staining method (DAKO) was performed, followed by counterstaining with hemalaun (Merck). Negative controls were performed by omitting the first antibody. The slides were evaluated using the light microscope BX53 (Olympus).
10
Protein Expression Analysis via SDS-PAGE
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The extracted samples were subjected to SDS-PAGE by using specific antibody for Bcl2 (1:1000), Bax (1:1000), Caspase3 (1:1000), GAPDH (1:1000) (Millipore, Germany). The bands were detected with an EZ-ECL kit (BI Biological Industries, 20-500-120) in Imagequant LAS4000 mini (GE Healthcare) after incubation with secondary antibodies. Plus Image-Pro 6.0 software was used for image analysis. GAPDH was used as control for normalization40 (link).
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!