The protein levels of aorta tissues and RAW264.7 macrophages were examined using western blot. The protein was extracted by lysing the aortic tissues and cells with lysis buffer supplemented with 1% protease inhibitors and phosphatase inhibitors (MedChemExpress, USA) on ice. After centrifugation, the supernatant was collected and quantified the concentration using a BCA kit (Boster Biological Technology, Wuhan, China). Then the equal amount of proteins (20 μg) were separated by 10%-12% SDS-PAGE and transferred to PVDF membranes (Millipore). After blocking with 5% non-fat milk, the membranes were probed with different primary antibodies overnight at 4℃, including PI3K, p-PI3K (Tyr 458), Akt, p-Akt (Ser 473), mTOR, p-mTOR (Ser 2448), ULK1, p-ULK1 (Ser 317), p70S6 kinase (S6K), p-S6K (Thr389), S6, p-S6 (Ser235/236), p62, iNOS (Cell Signaling Technology, Boston, USA), LC3 (Proteintech Biotechnology, Chicago, USA), Beclin1, ABCA1, ABCG1 (Abcam, Cambridge, UK), Arg-1, CD206, and GAPDH, β-actin (Abclonal, Boston, USA). Subsequently, the membranes were incubated with horseradish peroxidase-conjugated secondary antibodies (Boster Biological Technology, Wuhan, China). The proteins bands were visualized using ECL kit (Boster Biological Technology, Wuhan, China) and analyzed with Image J software (NIH, USA).
Protease inhibitors and phosphatase inhibitors
Manufactured by MedChemExpress
Sourced in United States
Protease inhibitors and phosphatase inhibitors are laboratory reagents used in various biochemical and cell biology applications. Protease inhibitors are used to prevent the degradation of proteins by proteolytic enzymes, while phosphatase inhibitors are used to maintain the phosphorylation state of proteins. These reagents are commonly used in protein purification, enzyme activity assays, and cell signaling studies.
Automatically generated - may contain errors
Lab products found in correlation
Pvdf membrane, by Merck Group (3 mentions)
Horseradish peroxidase conjugated secondary antibody, by Boster Bio (3 mentions)
Ecl kit, by Boster Bio (3 mentions)
Beclin 1, by Abcam (3 mentions)
Bca kit, by Boster Bio (3 mentions)
Cd206, by ABclonal (3 mentions)
Abca1, by Abcam (3 mentions)
Arg 1, by ABclonal (3 mentions)
P s6k, by Cell Signaling Technology (2 mentions)
P akt, by Cell Signaling Technology (1 mentions)
P ulk1, by Cell Signaling Technology (1 mentions)
Abcg1, by Abcam (1 mentions)
P mtor, by Cell Signaling Technology (1 mentions)
P pi3k, by Cell Signaling Technology (1 mentions)
3 protocols using protease inhibitors and phosphatase inhibitors
1
Western Blot Analysis of Aorta and Macrophages
2
Protein Expression Analysis in Aorta and Macrophages
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The protein levels of aorta tissues and RAW264.7 macrophages were examined using western blot. The protein was extracted by lysing the aortic tissues and cells with lysis buffer supplemented with 1% protease inhibitors and phosphatase inhibitors (MedChemExpress, USA) on ice. After centrifugation, the supernatant was collected and quantified the concentration using a BCA kit (Boster Biological Technology, Wuhan, China). Then the equal amount of proteins (20 µg) were separated by 10%-12% SDS-PAGE and transferred to PVDF membranes (Millipore). After blocking with 5% non-fat milk, the membranes were probed with different primary antibodies overnight at 4 , including PI3K, p-PI3K (Tyr 458), Akt, p-Akt (Ser 473), mTOR, p-mTOR (Ser 2448), ULK1, p-ULK1 (Ser 317), p62, iNOS (Cell Signaling Technology, Boston, USA), LC3 (Proteintech Biotechnology, Chicago, USA), Beclin1, ABCA1, ABCG1 (Abcam, Cambridge, UK), Arg-1, CD206, and GAPDH, β-actin (Abclonal, Boston, USA). Subsequently, the membranes were incubated with horseradish peroxidase-conjugated secondary antibodies (Boster Biological Technology, Wuhan, China). The proteins bands were visualized using ECL kit (Boster Biological Technology, Wuhan, China) and analyzed with Image J software (NIH, USA).
3
Western Blot Analysis of Aortic Tissue and Macrophages
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The protein levels of aorta tissues and RAW264.7 macrophages were examined using western blot. The protein was extracted by lysing the aortic tissues and cells with lysis buffer supplemented with 1 % protease inhibitors and phosphatase inhibitors (MedChemExpress, USA) on ice. After centrifugation, the supernatant was collected and quanti ed the concentration using a BCA kit (Boster Biological Technology, Wuhan, China). Then the equal amount of proteins (20 μg) were separated by 10 %-12 % SDS-PAGE and transferred to PVDF membranes (Millipore). After blocking with 5 % non-fat milk, the membranes were probed with different primary antibodies overnight at 4℃, including PI3K, p-PI3K (Tyr 458), Akt, p-Akt (Ser 473), mTOR, p-mTOR (Ser 2448), ULK1, p-ULK1 (Ser 317), p70S6 kinase (S6K), p-S6K (Thr389), S6, p-S6 (Ser235/236), p62, iNOS (Cell Signaling Technology, Boston, USA), LC3 (Proteintech Biotechnology, Chicago, USA), Beclin1, ABCA1, ABCG1 (Abcam, Cambridge, UK), Arg-1, CD206, and GAPDH, β-actin (Abclonal, Boston, USA). Subsequently, the membranes were incubated with horseradish peroxidase-conjugated secondary antibodies (Boster Biological Technology, Wuhan, China). The proteins bands were visualized using ECL kit (Boster Biological Technology, Wuhan, China) and analyzed with Image J software (NIH, USA).
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