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• Western blot analysis of recombinant Akt1, Akt2 and Akt3 proteins, and extracts from HeLa, C2C12, C6 and COS cells, using Akt (pan) (11E7) Rabbit mAb. • Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Akt (pan) (11E7) Rabbit mAb in the presence of control peptide (left) or Akt (pan) (11E7) Blocking Peptide #1085 (right). • Immunohistochemical analysis of paraffin-embedded human lung carcinoma, using Akt (pan) (11E7) Rabbit mAb. • Immunohistochemical analysis of paraffin-embedded human melanoma, using Akt (11E7) Rabbit mAb. • Immunohistochemical analysis using Akt (pan) (11E7) Rabbit mAb on SignalSlide(TM) Phospho-Akt (Ser473) IHC Controls #8101 (paraffin-embedded LNCaP cells, untreated (left) or LY294002-treated (right)). Note the lack of phosphorylated Akt-associated stain at the membrane of the LY294002 treated cells. • Immunohistochemical analysis of paraffin-embedded HeLa cells untreated (left) or transfected with Akt 1/2/3 Kinases ShortCut® siRNA Mix (New England BioLabs #N2005) (right), using Akt (pan) (11E7) Rabbit mAb (top) or Cleaved Caspase-3 (Asp175) #9661 (bottom).

Note the induction of cleaved caspase-3 in Akt deficient cells. • Confocal immunofluorescent analysis of HeLa cells, serum-starved (left) or insulin-treated (right), using Akt (pan) (11E7) Rabbit mAb (green). Blue pseudocolor = DRAQ5 ® #4084 (fluorescent DNA dye). • Flow cytometric analysis of untreated Jurkat cells, using Akt (pan) (11E7) Rabbit mAb (blue) compared to a nonspecific negative control antibody (red).

Western Blotting Protocol For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween ® 20 at 4°C with gentle shaking, overnight. NOTE: Please refer to primary antibody datasheet or product webpage for recommended antibody dilution. Solutions and Reagents From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: Western Blotting Application Solutions Kit NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.