Anti-Phospho-NF-kB p65 (Ser276) Polyclonal Antibody is a Rabbit antibody targeting Phospho-NF-kB p65 (Ser276). Anti-Phospho-NF-kB p65 (Ser276) Polyclonal Antibody can be used in IHC-P,IHC-Fr,ICC/IF,IF,FCM.

IgG

Human,Mouse,Rat (predicted:Dog,Pig,Cow,Horse)

1. Blank control (blue): Jurkat cells (fixed with 2% paraformaldehyde (10 min), then permeabilized with 90% ice-cold methanol for 30 min on ice).
Primary Antibody: Rabbit Anti-NFKB p65 (Ser276) antibody antibody (TMAB-01464), Dilution: 0.2 μg in 100 μL 1X PBS containing 0.5% BSA;
Isotype Control Antibody: Rabbit Igg (orange),used under the same conditions);
Secondary Antibody: Goat anti-rabbit IgG-Pe (white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA.
2. Paraformaldehyde-fixed, paraffin embedded (Rat urinary bladder); Antigen retrieval by microwave in sodium citrate buffer (pH6.0); Block endogenous peroxidase by 3% hydrogen peroxide for 30 minutes; Blocking buffer (3% BSA) at RT for 30 min; Antibody incubation with (Phospho-NFKB p65 (Ser276)) Polyclonal Antibody, Unconjugated (TMAB-01464) at 1:400 overnight at 4°C, followed by conjugation to the secondary antibody (labeled with HRP) and DAB staining.
3. Tissue/cell: Hela cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum) at 37°C for 20 min; Antibody incubation with (Phospho-NFKB p65 (Ser276)) polyclonal Antibody, Unconjugated (TMAB-01464) 1:100, 90 minutes at 37°C; followed by a FITC conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue) was used to stain the cell nucleus.
4. Tissue/cell: MCF7 cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum) at 37°C for 20 min; Antibody incubation with (Phospho-NFKB p65 (Ser276)) polyclonal Antibody,Unconjugated 1:100, 90 minutes at 37°C; followed by a FITC conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue) was used to stain the cell nucleus.
5. Blank control: A431. Primary Antibody (green line): Rabbit Anti-Phospho-NFKB p65 (Ser276) antibody (TMAB-01464)
Dilution: 1 μg/10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG.
Secondary Antibody: Goat anti-rabbit IgG-FITC
Dilution: 1 μg/test.
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°C. 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.
6. Blank control: HL-60. Primary Antibody (green line): Rabbit Anti-Phospho-NFKB p65 (Ser276) antibody (TMAB-01464)
Dilution: 2 μg/10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG.
Secondary Antibody: Goat anti-rabbit IgG-FITC
Dilution: 1 μg/test.
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°C. 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.

IHC-P: 1:100-500; IHC-Fr: 1:100-500; ICC/IF: 1:100; IF: 1:100-500; FCM: 0.2μg /Test

Polyclonal

KLH conjugated Synthesised phosphopeptide: human NFKBp65 around the phosphorylation site of Ser276

Human

RelA (p65),NFkB pathway,Inflammatory mediators,RelA (p65),NFkB pathway,Obesity,Host Immune Response,Host Virus Interaction,Alzheimer's disease,NFkB Pathway