Total proteins were isolated using RIPA buffer (Beyotime, Shanghai, China). Subsequently, proteins were separated by the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and then transferred onto the polyvinylidene difluoride (PVDF) membranes. After blocking with 5% non-fat milk for 2 h, the membranes were incubated with the primary antibodies against SHMT2 (1:1000, ab180786, Abcam, Cambridge, MA, USA), hexokinase-2 (HK-2) (1:5000, ab104836, Abcam), lactate dehydrogenase A chain (LDHA) (1:3000, ab135366, Abcam), glucose transporter 1 (GLUT1) (1:10,000, ab40084, Abcam), and then incubated with the secondary HRP-conjugated antibody (1:1000, ab9482, Abcam). Protein bands were visualized using ECL detection reagent and normalized by β-actin (1:1000, 4967, Cell Signaling Technology, Inc., Beverly, MA, USA). Triplicate individual experiments were performed in this study.
Ab135366
Manufactured by Abcam
Sourced in United States
Ab135366 is a laboratory equipment product. It is a tool used for scientific research and experimentation. The core function of this product is to provide a specific capability or feature to assist in the conduct of laboratory procedures and analysis. No further details about its intended use or specific capabilities can be provided in an unbiased and factual manner.
Automatically generated - may contain errors
Lab products found in correlation
Ab9482, by Abcam (3 mentions)
Ripa buffer, by Beyotime (2 mentions)
Ab180786, by Abcam (2 mentions)
Ab40084, by Abcam (2 mentions)
Ab104836, by Abcam (2 mentions)
β actin, by Cell Signaling Technology (2 mentions)
Polyvinylidene difluoride membrane, by Merck Group (1 mentions)
Ab174830, by Abcam (1 mentions)
Ab32150, by Abcam (1 mentions)
Chemiluminescence chromogenic substrate, by Beyotime (1 mentions)
Ab16667, by Abcam (1 mentions)
Ab2302, by Abcam (1 mentions)
Ab2732, by Abcam (1 mentions)
Ab40776, by Abcam (1 mentions)
Ab109268, by Abcam (1 mentions)
Ab182651, by Abcam (1 mentions)
3 protocols using ab135366
1
Western Blot Analysis of Metabolic Proteins
2
Protein Extraction and Western Blot Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Total protein was extracted from tissues and cells using radioimmunoprecipitation assay (RIPA) buffer. Then, isolated protein was separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and electrophoretically transferred to polyvinylidene difluoride membrane (Millipore, Billerica, MA, USA). After that, immunoblotting was conducted using primary antibodies against Ki67 (1:1,000, ab16667, Abcam, Cambridge, MA, USA), pro-caspase-3 (pro-c3) (1:1,000, ab32150, Abcam), cleaved-caspase-3 (cleaved-c3) (1:1,000, ab2302, Abcam), lactate dehydrogenase A chain (LDHA) (1:3,000, ab135366, Abcam), HMGCR (1:5,000, ab174830, Abcam), and HRP-conjugated secondary antibody (1:1,000, ab9482, Abcam). GAPDH was as used as an endogenous control. Finally, protein signals were visualized using the chemiluminescence chromogenic substrate (Beyotime).
3
Protein Expression Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Total proteins were isolated using RIPA buffer (Beyotime, Shanghai, China). Subsequently, proteins were separated by the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and then transferred onto the polyvinylidene di uoride (PVDF) membranes. After blocking with 5% non-fat milk for 2 h, the membranes were incubated with the primary antibodies against SHMT2 (1:1000, ab180786, Abcam, Cambridge, MA, USA), hexokinase-2 (HK-2) (1:5000, ab104836, Abcam), lactate dehydrogenase A chain (LDHA) (1:3000, ab135366, Abcam), glucose transporter 1 (GLUT1) (1:10000, ab40084, Abcam), phosphoinositide 3-kinase (PI3K) (1:1000, ab40776, Abcam), phosphorylated(p)-PI3K (p-PI3K) (1:1000, ab182651, Abcam), protein kinase B (AKT) (1:1,000, 9272, Cell Signaling Technology, Boston, MA, USA), p-AKT (1:1,000, 9271, Cell Signaling Technology), mammalian target of rapamycin (mTOR) (1:2000, ab2732, Abcam), p-mTOR (1:5000, ab109268, Abcam) and then interacted with the secondary HRP-conjugated antibody (1:1000, ab9482, Abcam). Protein bands were visualized using electrochemiluminescence and normalized by β-actin (1:1,000, 4967, Cell Signaling Technology).
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