Anti-Phospho-AKT3 (Tyr473) Polyclonal Antibody is a Rabbit antibody targeting Phospho-AKT3 (Tyr473). Anti-Phospho-AKT3 (Tyr473) Polyclonal Antibody can be used in WB,IHC-P,IHC-Fr,IF,FCM.

IgG

Human,Mouse,Rat

1. Paraformaldehyde-fixed, paraffin embedded (Rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-AKT3 (Tyr473)) Polyclonal Antibody, Unconjugated (TMAB-10302) at 1:400 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
2. Paraformaldehyde-fixed, paraffin embedded (Mouse brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-AKT3 (Tyr473)) Polyclonal Antibody, Unconjugated (TMAB-10302) at 1:400 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
3. Sample:
Small intestine (Mouse) Lysate at 40 μg
Large intestine (Mouse) Lysate at 40 μg
Primary: Anti-phospho-AKT3 (Tyr473) (TMAB-10302) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 57 kD
Observed band size: 57 kD
4. Blank control (black line): Hela.
Primary Antibody (green line): Rabbit Anti-phospho-AKT3 (Tyr473) antibody (TMAB-10302)
Dilution: 2 μg/Test;
Secondary Antibody (white blue line): Goat anti-rabbit IgG-FITC
Dilution: 0.5 μg/Test.
Isotype control (orange line): Normal Rabbit IgG
Protocol
The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 90% ice-cold methanol for 20 min at -20℃, The cells were then incubated in 5% BSA to block non-specific protein-protein interactions for 30 min at room temperature. Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.

WB: 1:500-2000; IHC-P: 1:100-500; IHC-Fr: 1:100-500; IF: 1:100-500; FCM: 2μg/Test

Polyclonal

KLH conjugated synthesised phosphopeptide: human AKT3 around the phosphorylation site of Tyr473

Human

AKT3

RAC-gamma serine/threonine-protein kinase