The penumbra tissue was treated with RIPA lysis buffer (Yamei, Suzhou), and the protein concentration was measured. 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was used to separate the target proteins, which were then transferred to a nitrocellulose membrane. The membrane was blocked with 5% BSA for 1 h and then incubated with 5% BSA containing corresponding primary antibodies, including JAK2 (YT2426, Immunoway, Plano, TX, USA) (1 : 400), p-JAK2 (3776, Cell Signaling Technology, Danvers, MA, USA) (1 : 500), STAT6 (YT4454, Immunoway) (1 : 400), p-STAT6 (YP0256, Immunoway) (1 : 400), Bcl-2 (YT0470, Immunoway) (1 : 500), Bax (YT0455, Immunoway) (1 : 500), and GAPDH (YM3029, Immunoway) (1 : 5,000) on a shaker at 4°C overnight. On the second day, Tris-buffered saline-tween (TBST) was used to rinse the membranes 3 times, which were then incubated with secondary antibodies (RS23710, RS23920, Immunoway) (1 : 10,000) for 1 h. The bands were observed using an Odyssey Infrared Imaging System 3.0.29 (LICOR, Nebraska, USA).
Bcl 2
Manufactured by Immunoway
Sourced in United States, China
Bcl-2 is a protein that plays a crucial role in regulating apoptosis, or programmed cell death. It functions as an anti-apoptotic protein, helping to prevent cells from undergoing apoptosis. Bcl-2 is an important component in the study of cellular processes and pathways related to cell survival and death.
Automatically generated - may contain errors
Lab products found in correlation
Bcl2 associated x (bax), by Immunoway (11 mentions)
Caspase3, by Immunoway (4 mentions)
Gapdh, by Immunoway (3 mentions)
β actin, by Immunoway (3 mentions)
Vascular endothelial growth factor (vegf), by Immunoway (3 mentions)
Hif 1α, by Immunoway (2 mentions)
Cleaved caspase3, by Immunoway (2 mentions)
Ym3029, by Immunoway (1 mentions)
P jak2, by Cell Signaling Technology (1 mentions)
P stat6, by Immunoway (1 mentions)
Rs23710, by Immunoway (1 mentions)
Rs23920, by Immunoway (1 mentions)
Odyssey infrared imaging system 3, by LI COR (1 mentions)
P akt, by Abcam (1 mentions)
5 lox, by Abcam (1 mentions)
Nf κb, by Immunoway (1 mentions)
Gapdh, by Abcam (1 mentions)
Bcl2 associated x (bax), by Abcam (1 mentions)
Ab32042, by Abcam (1 mentions)
Ptpn1, by Abcam (1 mentions)
15 protocols using bcl 2
1
Western Blot Analysis of JAK2/STAT6 Signaling
2
Evaluating Astragalus' Apoptosis Mechanisms
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Astragalus roots were purchased from Tianjin Tongrentang Group Co., Ltd. (Tianjin, China). Bax, β-actin, Bcl-2, and Cyt c antibodies were purchased from ImmunoWay Biotechnology Company (Plano, TX, USA). Apoptosis index-related detection kits for cell experiments were all provided by Beijing Solarbio Science & Technology Co., Ltd. (Beijing, China). Human hepatoma HepG2 cells were provided by the Shanghai Institute of Biological Sciences at the Chinese Academy of Sciences (Shanghai, China). All other chemicals used in this study were of analytical grade.
3
Protein Expression Analysis After SAH
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Brain tissues were collected at different time points after SAH induction , Western blot was performed as previously described [12 (link)]. Primary antibodies used included 5-LOX (1:5000, Abcam), p-AKT (1:5000, Abcam), AKT (1:5000, Abcam), Bax (1:5000, Abcam), GAPDH (1:5000, Abcam), NF-κB (1:1000, Immunoway), Bcl2 (1:1000, Immunoway), and histone (1:1000, Proteintech). Blots were detected by enhanced chemiluminescence and quantified using Quantity One software. GAPDH and histone were used as loading controls for whole cell and nuclear proteins.
4
Western Blot Analysis of Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total protein was extracted from cells after transfection using lysis buffer for western and IP (KeyGen, KGP701, Nanjing, China), and protein concentration was measured using a BCA protein assay kit (Beyotime, P0010, Shanghai, China). Proteinsamples were electrophoresed using SDS-PAGE and then transferred for incubation at 4ºC overnight with the following specific primary antibodies: p53 (ImmunoWay, YT3528, TX, USA), Bax (ImmunoWay, YT0455, TX, USA), Bcl-2 (ImmunoWay, YM3041, TX, USA), active caspase-3 (Abcam, ab32042, Cambridge, UK), PTPN1 (Abcam, ab75856, Cambridge, UK), MAP3K11 (Abcam, ab51068, Cambridge, UK), JNK1/2/3 (Bimake, A5005, TX, USA), c-Jun (Bimake, A5730, TX, USA), and GAPDH (ImmunoWay, YM3445, TX, USA). The samples were then incubated with HRP-conjugated anti-rabbit IgG antibody (CST, #7074, MA, USA) at room temperature for 1 h. Finally, proteins were detected using ECL (KeyGEN, KGP902, Nanjing, China). GAPDH was used as an internal reference. Image J was used for quantification of all Western blot bands.
5
In Vitro Evaluation of Antioxidant and Antiapoptotic Effects of Traditional Chinese Medicine
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All herbal medicines were provided by Guangyuan Hospital of Traditional Chinese Medicine (Guangyuan, China). Fetal bovine serum (FBS) and Dulbecco’s modified Eagle medium (DMEM) were purchased from the Gibco Co. (Grand Island, NY, United States). BCA protein assay reagents, SDS-polyacrylamide gel electrophoresis (SDS-PAGE) preparation kit, penicillin-streptomycin mixture, trypsin, phosphate buffer saline (PBS) and cell counting kit-8 (CCK-8) were purchased from Boster Biological Technology Co., Ltd. (Wuhan, China). Annexin V-FITC/PI assay kit was purchased from Multisciences (Lianke) Biotechnology Corporate Limited (Hangzhou, China). H2O2 was purchased from Chengdu Chron Chemicals Co. Ltd. (Chengdu, China). Primary antibodies for PI3K, phosphorylation-PI3K (p-PI3K), AKT, phosphorylation-AKT (p-AKT), Nrf2, Bcl-2, Bax and cleaved-caspase-3 (C- caspase-3) were obtained from the ImmunoWay Biotechnology Co. (Suzhou, China). The assay kits for MDA, SOD, CAT, and GSH-PX were purchased from the Nanjing Jiancheng Bioengineering Institute (Nanjing, China). Ultrapure water purified by Millipore Ultra-pure Water Purifier (Millipore, Milford, MA, United States) was used. Other reagents were all of analytical grade.
6
Protein Expression Analysis in Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Proteins were extracted from samples (tissue and cultured cells) using RIPA buffer containing 1 mM phenylmethanesulfonyl fluoride (PMSF) (SolarBio) and separated by SDS-PAGE at 8–10%. The proteins were then transferred to a polyvinylidene fluoride membrane to be probed with Bax (Immunoway, 1:1,000), Bcl-2 (Immunoway, 1:1,000), Caspase 3 (Immunoway, 1:1,000), MMP2 (Immunoway, 1:1,000), Wnt3 (Immunoway, 1:1,000), Wnt5a (Immunoway, 1:1,000), β-catenin (Proteintech, 1:1,000), TCF-4 (Proteintech, 1:1,000), LEF-1(Proteintech, 1:1,000), IL-8 (Immunoway, 1:1,000) and GAPDH (Proteintech, 1:1,000) for 24 h at 4 ℃. Next, the proteins were incubated with appropriate secondary antibody for 2 h at approximately 25 ℃. All western blots were repeated at least 3 times. The density of the immunoreactive bands was quantified using ImageJ software.
7
Western Blot Analysis of Apoptosis Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Proteins from cell lysates or tissue lysates were separated by a 10% SDS polyacrylamide gel electrophoresis and transferred to a polyvinylidene fluoride membrane. After being blocked in 5% nonfat milk, protein blots were probed with a primary antibody followed by incubation with a peroxidase-conjugated secondary antibody. The primary antibodies included HIF-1α (1:1000; ImmunoWay), Bax (1:1000; ImmunoWay), Bcl-2 (1:1000; ImmunoWay), caspase3 (1:1000; ImmunoWay), cleaved-caspase3 (1:1000; ImmunoWay), and PTEN (1:500; ImmunoWay). Chemiluminescence was detected by the ECL-plus kit (Beyotime). Band intensity was quantified by Image J software.
8
Quantitative Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Proteins were separated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and then transferred to a nitrocellulose filter membrane (NC, Pall Corporation, Pensacola, FL, USA). The membranes were blocked with 5% non-fatty milk and then probed with antibodies against RNF2 (1:500, Abcam, Cambridge, MA, USA), PIM3 (1:500, Abcam, Cambridge, MA, USA), HAX1 (1:500, Santa Cruz Biotechnology, Santa Cruz, CA, USA), AKT1 (1:1000, Cell Signaling Technology, Beverly, MA, USA), pAKT1 (1:1000, Cell Signaling Technology, Beverly, MA, USA), BCL2 (1:500, Immunoway, Newark, DE, USA) and β-actin (1:100000, Sigma, Louis, MO, USA). After washing with PBST (pH 7.4), the blots were incubated with horseradish peroxidase-conjugated secondary antibodies (1:2000, Zhongshan, Beijing, China). An ECL kit (Millipore, Billerica, MA, USA) was used to detect the intensity of each protein.
9
Oxidative Stress-Induced Neuronal Apoptosis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Reagents include lentivirus plasmid (Cyagen Biosciences), potassium ferrocyanide (Sigma-Aldrich), DAPI (2 mg/ml, Servicebio), DHE (Cayman Chemical), protease inhibitors (Thermo Fisher), and phosphatase inhibitors (Servicebio). The TUNEL kit was purchased from Vazyme Biotech. The BCA protein assay kit was purchased from CoWin Biosciences. RNA extraction kit was purchased from Tiangen Biotech.
Antibodies used were as follows: hepcidin (1 : 100, Affinity), TfR1 (1 : 10000, Invitrogen), FPN1 (1 : 5000 for WB, 1 : 200 for IF, Alpha Diagnostic International), FTL (1 : 5000 for WB, 1 : 200 for IF, Abcam), GFAP (1 : 100, Servicebio), NeuN (1 : 200, Abcam), Bcl-2 (1 : 2000, ImmunoWay), Bax (1 : 1000, Servicebio), p-JNK (1 : 1000, Cell Signaling Technology), JNK (1 : 1000, Arigo Biolaboratories), NOX2 (1 : 2000, GeneTex), 4-HNE (1 : 200 for IF, 1 : 1000 for WB, Arigo Biolaboratories), BDNF (1 : 1000, Servicebio), β-actin (1 : 1000, Cwbiotech), and β-tubulin (1 : 1000, Servicebio).
Antibodies used were as follows: hepcidin (1 : 100, Affinity), TfR1 (1 : 10000, Invitrogen), FPN1 (1 : 5000 for WB, 1 : 200 for IF, Alpha Diagnostic International), FTL (1 : 5000 for WB, 1 : 200 for IF, Abcam), GFAP (1 : 100, Servicebio), NeuN (1 : 200, Abcam), Bcl-2 (1 : 2000, ImmunoWay), Bax (1 : 1000, Servicebio), p-JNK (1 : 1000, Cell Signaling Technology), JNK (1 : 1000, Arigo Biolaboratories), NOX2 (1 : 2000, GeneTex), 4-HNE (1 : 200 for IF, 1 : 1000 for WB, Arigo Biolaboratories), BDNF (1 : 1000, Servicebio), β-actin (1 : 1000, Cwbiotech), and β-tubulin (1 : 1000, Servicebio).
10
Western Blot Analysis of Apoptosis and Autophagy
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Immediately after the animals were sacrificed, the flap tissue of area II was collected and analyzed by Western blot. For Western blot analysis, each sample was treated with lysis buffer to release tissue proteins and the protein concentration was measured by a BCA method. Next, gel electrophoresis was used to separate the proteins, which were then electrotransferred to PVDF membranes and blocked with nonfat milk 5%. The samples were eluted with TBST three times and incubated at 4 °C after adding the primary antibody solution. The incubation concentration of the primary antibody is as follows: Bax (1:1000, Immunoway), Bcl-2 (1:1000, Immunoway), β-actin (1:5000, Immunoway), VEGF (1:1000, Immunoway), Nrf2 (1:1000, Immunoway), LC3B(1:2000, Abcam), SQSTM1/p62 (1:30,000, Abcam), CTSD (1:5000, Abcam), GAPDH (1:5000, Immunoway). Subsequently, secondary antibodies conjugated to HRP were incubated with the samples for 2 h at room temperature. Finally, an ECL kit was used to visualize the bands on the membrane, and the intensity of the bands was quantified using Image Lab software.
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!