Dissected hippocampal tissues were homogenized in RIPA buffer (1% NP‐40, 0.1% SDS, 0.5% PMSF, and 4% Roche protease inhibitor) by sonication. After centrifugating at 12,000g for 20 min at 4°C, the supernatant was collected and protein concentration was measured. A total cell extract containing ~30 μg of protein was subjected to SDS‐PAGE and then transferred onto NC membrane. The membrane was blocked by incubation with 5% skim milk for 90 min and then hybridized overnight with primary antibodies at 4°C. The primary antibodies used include anti‐ferritin light chain (ab109373, Abcam), anti‐ferritin heavy chain (ab183781, Abcam), anti‐TfR1 (13–6800, Invitrogen), anti‐Bcl2 (12789‐1‐AP, Proteintech), anti‐Bax (50599‐2‐Ig, Proteintech), anti‐brain‐derived neurotrophic factor (BDNF) (ab108319, Abcam), anti‐furin (ab183495, Abcam), and anti‐β‐actin (CW0096, CWBIO). After washing with tris‐buffer saline with 0.05% Tween‐20, the membrane was incubated at room temperature for 90 min with anti‐rabbit or anti‐mouse secondary antibody conjugated with horseradish peroxide (Zhongshan Biotechnology). Immunoreactive bands were detected by the enhanced chemiluminescence detection kit (Amersham, UK) and visualized by a Bio‐Rad chemiluminescence imager.
Anti bcl2 12789 1 ap
Manufactured by Proteintech
Sourced in United States
Anti-Bcl2 (12789-1-AP) is an antibody product offered by Proteintech. It is designed to detect the Bcl2 protein, which is involved in the regulation of apoptosis, or programmed cell death. The antibody can be used in various research applications, such as Western blotting, immunohistochemistry, and immunofluorescence, to study the expression and localization of Bcl2 in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Anti gapdh 60004 1 ig, by Proteintech (2 mentions)
Anti beclin 1, by Cell Signaling Technology (2 mentions)
Anti β catenin, by Cell Signaling Technology (2 mentions)
Anti tcf4, by Cell Signaling Technology (2 mentions)
Enhanced chemiluminescence detection kit, by Cytiva (1 mentions)
Cw0096, by CWBIO (1 mentions)
Chemiluminescence imager, by Bio-Rad (1 mentions)
Ab109373, by Abcam (1 mentions)
Ab183781, by Abcam (1 mentions)
Ab183495, by Abcam (1 mentions)
Ab108319, by Abcam (1 mentions)
Protein a g agarose, by Merck Group (1 mentions)
Anti cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Anti cleaved parp, by Cell Signaling Technology (1 mentions)
Epidermal growth factor (egf), by Merck Group (1 mentions)
Anti akt, by Cell Signaling Technology (1 mentions)
Anti c cbl, by Cell Signaling Technology (1 mentions)
Ag1478, by Selleck Chemicals (1 mentions)
Alexa flour 488 conjugated anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
Anti egfr, by Cell Signaling Technology (1 mentions)
9 protocols using anti bcl2 12789 1 ap
1
Hippocampal Protein Expression Analysis
2
Antibody and Reagent Specifications for Cell Signaling Experiments
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Puromycin and G-418 were purchased from Cayman Chemical (Ann Arbor, Michigan, USA). AG1478, and U0126 were purchased from Selleck. The protein A/G agarose and EGF were purchased from Sigma (St louis, MO, USA). The following antibodies were used: anti-β-actin (#8457), anti-cleaved-caspase-3 (#9664), ani-caspase-3 (#9662), anti-cleaved PARP (#5625), anti-EGFR (#4267), anti-Erk1/2 (#4695), anti-phospho-Erk1/2 (#4370), anti-p38 (#8690), anti-phospho-p38 (#9211), anti-AKT(#9272), anti-phospho-AKT (Ser473) (#4060), anti-phospho-JNK (Thr183/Tyr185) (#9251), anti-JNK (#9252), anti-HA-Tag (#3724), anti-c-Cbl (#2747), anti-ubiquitin (P4D1) (#3936) from Cell Signaling Technology (Beverly, MA, USA); anti-Flag-Tag from Sigma-Aldrich (St Louis, MO, USA), anti-GAPDH (AC002), anti-phospho-EGFR (Y1068) (AP0820) from Abclonal Technology(Shanghai, China); anti-14-3-3σ (ab14123), anti-Bim (ab7888) from Abcam (Cambridge, MA, USA); anti-Bcl-2 (12789-1-AP) from proteintech (Chicago, IL, USA); Alexa Flour 488-conjugated anti- rabbit IgG, Alexa Flour 555-conjugated anti-rabbit IgG and Alexa Flour 555-conjugated anti-mouse IgG from Thermo Scientific (Rockford, IL, USA). HRP conjugated anti-rabbit IgG, HRP conjugated anti-mouse IgG from Beyotime Institute of Biotechnology; APC anti-human EGFR antibody (#352905) used for cell surface immunofluorescence staining from BioLegend (San Diego, CA).
3
Protein Expression Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After RIPA cleavage, we extracted total protein and measured with BCA method. After quantitative denaturation, protein electrophoresis membrane transfer and blocked. The primary antibody anti-Bcl2 (12,789-1-AP, Proteintech), anti-Bax (50,599-2-Ig, Proteintech), anti-cleaved-caspases3 (ab2302, abcam), anti-DKK1 (21,112-1-AP, Proteintech), anti-p-GSK3β (#3548, Cell Signaling Technology), anti-GSK3β (#12,456, Cell Signaling Technology), anti-β-catenin (51,067-2-AP, Proteintech), Wnt1 (27,935-1-AP, proteintech), and anti-GAPDH (60,004-1-Ig, Proteintech) incubation and second incubation were carried out according to the operation steps. The expression of the protein was expressed by the gray value.
4
Protein Expression Analysis in Cell Lines
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total protein was extracted from cell lines using RIPA Lysis Buffer (Biosharp, China) containing a protease inhibitor and quantified using a BCA Protein Assay Kit (Biosharp, China) according to the manufacturer’s instructions. Western blotting was performed using a standard protocol. The primary antibodies were: anti-INPP4B (BM4609, Boster, USA), anti-PD-L1 (66248-1-lg, Proteintech), anti-p85 (660225-1-lg, Proteintech), anti-Phospho-AKT (66444–1-lg, Proteintech), anti-Bcl-2 (12789–1-AP, Proteintech), and anti-Cyclin D1 (60186–1-lg, Proteintech). In addition, an anti-GAPDH (60004–1-lg, Proteintech) was used as loading control.
5
Western Blot Analysis of Key Cellular Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell protein lysates were generated using RIPA Lysis Buffer (Beyotime, China). Cellular proteins were extracted 72 h after transfection. Western blotting was performed as reported previously 45 (link). Five μg proteins were boiled in 5 × SDS buffer for 5 min, separated by SDS-PAGE, and transferred to PVDF membranes (Millipore, USA). Then, the membranes were blocked with skim milk and probed with Anti-p53 (A0263, abclonal, USA), Anti-NFAT5 (Bs-9473R, bioss,China), Anti-β-catenin (Ab32572, Abcam, USA), Anti-c-myc (Ab32072, Abcam, USA), Anti-Bcl-2 (12789-1-ap, Proteintech, China). β-actin (ab8226, Santa Cruz, USA) was used as a loading control. The results were visualized with horseradish peroxidase-conjugated secondary antibodies (Santa Cruz) and enhanced chemiluminescence. All blots were performed in triplicate and protein expression levels were quantified relatively to the expression of β-actin using the Image J 1.42q software (Wayne Rasband).
6
Protein Expression Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The primary antibodies used were as follows: anti‐TRAF6 [8028Sl; Cell Signaling Technology (CST), MA, USA], anti‐ATG16L2 (1:500, D153556; Sangon Biotech, Shanghai, China), anti‐cleaved PARP (9542S; CST), anti‐cleaved Caspase‐9 (9508S; CST), anti‐BCL2 (12789‐1‐AP; Proteintech, IL, USA), anti‐Bax (5023S; CST), anti‐p62(D199564; Sangon Biotech), anti‐LC3 A/B (4108S; CST), anti‐c‐Jun (9165S; CST), anti‐Lamin A/C (sc‐376248; Santa Cruz, CA, USA), and anti‐GAPDH (60004‐1‐Ig; Proteintech).
7
Lung Cancer Cell Culture Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell culture. The NSCLC cell lines A549, H1299, H292, H460, and HCC827 and the normal human lung cell line BEAS-2B were obtained from the American Type Culture Collection (Manassas, VA, USA). The cell lines were grown in RPMI-1640 medium (HyClone, South Logan, VT, USA), supplemented with 10% (v/v) fetal bovine serum (Gibco, Grand Island, NY, USA) and 100 IU/ml penicillin and 100 µg/ml streptomycin (Beyotime Biotechnology, Shanghai, China). All of the cells were cultured at 37˚C in a humidified incubator with 5% CO 2 .
Reagents and antibodies. Cisplatin and Z-VAD-FMK were purchased from Selleck (Houston, TX, USA). SBI0206965 was obtained from DC Chemicals (Shanghai, China). The anti-Ulk1 (8054) and anti-LC3 (3868) antibodies were obtained from Cell Signaling Technology (Danvers, MA, USA). The anti-Bcl2 (12789-1-AP) and Bclxl (10783-1-AP) antibodies were purchased from Proteintech (Chicago, IL, USA). The anti-p62 (ab109012) antibody was purchased from Abcam (Cambridge, UK). The mouse anti-β-actin monoclonal antibody (AM1021B) was from Abgent (San Diego, CA, USA).
Reagents and antibodies. Cisplatin and Z-VAD-FMK were purchased from Selleck (Houston, TX, USA). SBI0206965 was obtained from DC Chemicals (Shanghai, China). The anti-Ulk1 (8054) and anti-LC3 (3868) antibodies were obtained from Cell Signaling Technology (Danvers, MA, USA). The anti-Bcl2 (12789-1-AP) and Bclxl (10783-1-AP) antibodies were purchased from Proteintech (Chicago, IL, USA). The anti-p62 (ab109012) antibody was purchased from Abcam (Cambridge, UK). The mouse anti-β-actin monoclonal antibody (AM1021B) was from Abgent (San Diego, CA, USA).
8
Western Blot Analysis of Key Cellular Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells or tissues were lysed with RIPA buffer (Beyotime) containing protease inhibitors cocktail (Selleck). Protein samples were loaded onto and separated using SDS-PAGE, then transferred onto polyvinylidene fluoride membranes (Milliore). The membranes were blocked for 1 h at room temperature and incubated with the anti-LC3B (#3868, Cell Signaling Technology), anti-ACTB (#3700, Cell Signaling Technology), anti-TCF4 (#2565, Cell Signaling Technology), anti-β-catenin (#8480, Cell Signaling Technology), anti-Bcl2 (12789-1-AP, Proteintech), or anti-Beclin1 (#3495, Cell Signaling Technology) antibodies overnight at 4 °C. After washing, the blots were incubated with goat anti-rabbit (111-035-003, Jackson) or anti-mouse (115-035-003, Jackson) horse radish peroxidase-conjugated secondary antibodies and visualized using the Immobilon™ horse radish peroxidase substrate peroxid solution (Millipore).
9
Co-Immunoprecipitation of key proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were lysed with immunoprecipitation (IP) lysis buffer (Beyotime). Total protein was incubated with 50 μl of Protein G Sepharose (GE healthcare) for 1 h at 4 °C on a rocking platform to reduce non-specific binding. After removing the beads, the supernatant was incubated with 5 μg anti-TCF4 (#2565, Cell Signaling Technology), anti-βcatenin (#8480, Cell Signaling Technology), anti-Bcl2 (12789-1-AP, Proteintech), anti-Beclin1 (#3495, Cell Signaling Technology) antibodies, or negative control antirabbit IgG (#2729, Cell Signaling Technology) overnight at 4 ℃. A total of 50 μl of Protein G Sepharose was added to each immunoprecipitation mixture, and the incubation was continued for 1 h at 4 °C on a rocking platform. The immunoprecipitates were collected and washed three times with the cold IP lysis buffer. After the loading buffer was added, the sepharose was boiled and subjected to western blot assay. The clean-blot IP detection reagent (horse radish peroxidase) secondary antibodies (21232, Thermo) were employed to avoid the denatured heavy and light chains from antibodies used in co-IP assays.
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!