Western blot analysis was performed on whole cell extracts (20 ug lysate) of HeLa (lane 1), L6 (lane 2), K562 (lane 3), Jurkat (lane 4), MCF7 (lane 5), Hep G2 (lane 6), HT-29 (lane 7), HCT 116 (lane 8) and MDA-MB-231 (lane 9). The blots were probed with Anti-AMPK alpha Mouse Monoclonal Antibody (Product# AHO1332, 0.5-2 ug/ml) and detected by chemiluminescence Goat anti-Mouse IgG (H+L) Secondary Antibody, HRP conjugate (Product # 626520, 1:4000 dilution). A 60 kDa band corresponding to AMPK alpha was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 12 % Bis-Tris gel (Product # NP0342BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
|Tested species reactivity||Human, Mouse, Rat|
|Host / Isotype||Mouse / IgG1, kappa|
|Immunogen||Recombinant fragment of human AMPK-alpha-1 expressed in E. coli.|
|Storage buffer||PBS, pH 7.2, with 1% BSA|
|Contains||0.1% sodium azide|
|Storage Conditions||Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.|
|Tested Applications||Dilution *|
|Western Blot (WB)||0.5-2 ug/ml|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
The protein encoded by this gene belongs to the ser/thr protein kinase family. It is the catalytic subunit of the 5'-prime-AMP-activated protein kinase (AMPK). AMPK is a cellular energy sensor conserved in all eukaryotic cells. The kinase activity of AMPK is activated by the stimuli that increase the cellular AMP/ATP ratio. AMPK regulates the activities of a number of key metabolic enzymes through phosphorylation. It protects cells from stresses that cause ATP depletion by switching off ATP-consuming biosynthetic pathways. Alternatively spliced transcript variants encoding distinct isoforms have been observed.
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