Human NSCLC tissues and adjacent non-cancerous tissues were cut into 4 µm slices. After baking at 70 °C for 30 minutes, the slices were dewaxed and hydrated using a xylene and alcohol series, respectively. Endogenous peroxidase activity was blocked by incubation with 3% hydrogen peroxide at room temperature for 10 minutes. Antigen recovery was performed by heating with a pressure cooker in citric acid antigen repair solution (pH 6.0). After blocking with 10% goat serum, the slices were incubated with rabbit anti-PSMD14 monoclonal antibody (ab109123) or USP14 polyclonal antibody (ab71165; Abcam, Cambridge, UK) at 4 °C overnight. The tissue sections were then washed in phosphate-buffered saline (PBS) and incubated with a horseradish peroxidase-conjugated secondary antibody (MaixinBiol, Fuzhou, China) for 1 hour at room temperature. Samples were stained with diaminobenzidine and counterstained with hematoxylin according to the MaxVision kit (MaixinBiol) instructions.
Ab71165
Manufactured by Abcam
Sourced in United States
Ab71165 is a polyclonal antibody produced in rabbit and directed against human Bcl-2 protein. Bcl-2 is a key regulator of apoptosis or programmed cell death.
Automatically generated - may contain errors
Lab products found in correlation
Ab109123, by Abcam (1 mentions)
Ab220162, by Abcam (1 mentions)
Dynabeads protein g magnetic beads, by Thermo Fisher Scientific (1 mentions)
Protease inhibitor cocktail, by Roche (1 mentions)
Ab230330, by Abcam (1 mentions)
Sds page gel, by Ipsen (1 mentions)
Pvdf 0.45 μm, by Merck Group (1 mentions)
Supersignal west dura extended duration substrate, by Thermo Fisher Scientific (1 mentions)
Anti β actin, by Proteintech (1 mentions)
Radioimmunoprecipitation assay lysis buffer, by Beyotime (1 mentions)
3 protocols using ab71165
1
Immunohistochemical Analysis of PSMD14 and USP14 in NSCLC
2
Co-Immunoprecipitation of USP14 and YTHDF1
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
BGC-823 cells were washed with PBS and incubated in 200 μL lysis buffer (50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 15 mM MgCl2, 5 mM EDTA, and 0.1% NP-40) containing protease inhibitor Cocktail (Roche, Mannheim, Germany) for subsequent Co-IP. The cell lysates were incubated with specific antibodies against USP14 (ab71165, Abcam, United States) and YTHDF1 (ab220162, Abcam, United States) at 4°C for 2 h, then incubated with 2.5 mg Dynabeads Protein G magnetic beads (Thermo Fisher Scientific, 10004D) at 4°C overnight. The beads were separated and washed with cold PBS, and Western blotting analysis was performed.
3
Western Blot Analysis of YTHDF1 and USP14
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
GC cells were obtained and extracted using Radio Immunoprecipitation Assay Lysis buffer (Beyotime, Shanghai, China) and equal volumes of GC cells extracted solution were separated on 7.5 and 10% SDS-PAGE gels (EpiZyme, Shanghai, China). The protein bands were transferred from SDS-PAGE gels to PVDF membranes (Millipore PVDF 0.45 μm). The primary antibodies anti-YTHDF1 (ab230330, Abcam, United States), anti-USP14 (ab71165, Abcam, United States), and anti-β-actin (Proteintech, IL, United States) were added at a dilution of 1:500 or 1:1,000 according to the manufacturer’s instructions and incubated at 4°C overnight. The secondary antibodies (horseradish peroxidase [HRP] rabbit IgG) were diluted 1:3,000 and incubated with the membranes at room temperature for 2 h. The membranes were washed thrice with Tris-Buffered Saline with Tween-20, and the immunoreactive bands were developed using SuperSignal West Dura Extended Duration Substrate (Thermo Fisher Scientific, IL, United States) based on established protocols. These experiments were repeated more than three times.
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!