Colon tissues or cells were lysed using RIPA extraction buffer (R0010, Solarbio, China). Protein concentrations were quantified using BCA protein assay kit (PC0020, Solarbio, China). Proteins were separated on 10% SDS polyacrylamide gels and then transferred to PVDF membranes. Membranes were blocked with quickblock solution (P0231, Beyotime, China) for 15 minutes, and then blocked with antibodies against STAT3 (4904, CST), p-STAT3 (9145, CST), Il-10 (A12255, ABclonal) and Tlr2 (13744, CST) at 4°C overnight. The membranes were then incubated with secondary antibody-conjugated HRP (1:10000, Abcam) for 1 hour at room temperature, and bands were visualized using an ECL kit (FD8000, Fdbio science, China). β-actin served as a loading control.
Ripa extraction buffer
Manufactured by Solarbio
Sourced in China
RIPA extraction buffer is a commonly used protein extraction and lysis buffer. It is designed to extract and solubilize cellular proteins for further analysis.
Automatically generated - may contain errors
Lab products found in correlation
Bca protein assay kit, by Solarbio (3 mentions)
Ecl kit, by Fdbio (2 mentions)
Hrp conjugated secondary antibody, by Abcam (1 mentions)
Quickblock solution, by Beyotime (1 mentions)
Cyp1a1, by Cell Signaling Technology (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Ecl kit, by Solarbio (1 mentions)
E cadherin, by Cell Signaling Technology (1 mentions)
Occludin, by Cell Signaling Technology (1 mentions)
Ab167161, by Abcam (1 mentions)
Et1601 4, by Huabio (1 mentions)
P21 sc 6246, by Santa Cruz Biotechnology (1 mentions)
Et1701 80, by Huabio (1 mentions)
Bca protein assay kit, by Meilun (1 mentions)
4 protocols using ripa extraction buffer
1
Western Blot Analysis of Colon Proteins
2
Protein Expression Analysis in Colonic Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The colonic tissues were lysed with RIPA extraction buffer (Solarbio, China) containing a 1% protease inhibitor cocktail. Following centrifugation at 12,000 rpm for 15 min at 4°C, the supernatant was collected and the total protein concentration was quantified using a bicinchoninic acid (BCA) protein assay kit (Solarbio, China). Proteins were separated by SDS–polyacrylamide gel electrophoresis and transferred onto polyvinylidene difluoride (PVDF) membranes (Millipore, USA). The membranes were blocked with 5% skim milk for 1.5 h and incubated with primary antibodies against PXR (Abcam, USA), E-cadherin (Cell Signaling Technology, USA), Cyp1a1 (Cell Signaling Technology, USA), and occludin (Cell Signaling Technology, USA) overnight at 4°C. The membranes were washed and incubated with HRP-labeled secondary antibodies at room temperature for 2 h, followed by signal detection with an ECL Kit (Solarbio, China).
3
Western Blot Analysis of Tight Junction Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Colon tissue was weighed and homogenized in RIPA extraction buffer (Solarbio, China). The homogenate was centrifuged at 4°C for 15 min at 15,000 g, then the supernatant was collected. The protein concentration was quantified with BCA protein assay kits (Solarbio, China) according to the manufacturer’s instructions. Proteins were separated by 10% SDS polyacrylamide gel, and then transferred onto PVDF membranes. The membranes were blocked with 5% skimmed milk for 1 h and then immunoblotted with primary antibodies against occuludin (ab167161, Abcam), claudin (36–4800, Thermo Fisher Scientific), helios (#89270, CST) at 4°C overnight. Membranes were then incubated with second antibodies labeled with HRP at room temperature for 1 h and bands were visualized using an ECL kits (Fdbio science, China). β-actin was used as a reference gene.
4
Western Blot Analysis of Stem Cell Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Tissue was weighed and homogenized in RIPA extraction buffer (Solarbio, China). The homogenate was centrifuged at 4 °C for 15 min at 15,000 × g, then the supernatant was collected. The protein concentration was quantified with BCA protein assay kits (Meilun, China) according to the manufacturer’s instructions. Proteins were separated by 10% SDS polyacrylamide gel and then transferred onto PVDF membranes. The membranes were blocked with 5% skimmed milk for 1 h and then immunoblotted with primary antibodies: p53 (ab167161, Abcam,1:1000), p21 (sc-6246,Santa Cruz,1:500), Lgr5 (Bioss, bs-20747R, 1:1000), c-Myc (CST, #5605S, 1:1000), Cyclin D1 (CST, # 2978, 1:1000), Active β-Catenin (CST. # 8814, 1:1000), Reg4 (Abclonal, A13129, 1:1000), β-actin (ET1701-80, HUABIO, 1:1000), Gapdh (ET1601-4, HUABIO,1:1000) at 4 °C overnight. Membranes were then incubated with second antibodies (HA1019, HA1006, HUABIO,1:10,000) labeled with HRP at room temperature for 2 h, and bands were visualized using an ECL kit (Fdbio science, China). β-Actin or Gapdh was used as a reference protein.
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!