Phospho-Smad2 (Ser465 + Ser467) Rabbit Polyclonal Antibody

• Catalog Number: orb6979
• Category: Antibodies
• Description: Phospho-Smad2 (Ser465 + Ser467) Rabbit Polyclonal Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: FC, ICC, IF, IHC-Fr, IHC-P, WB
• Predicted Reactivity: Bovine, Canine, Equine, Gallus, Porcine
• Reactivity: Human, Mouse, Rat
• Isotype: IgG
• Immunogen: KLH conjugated Synthesised phosphopeptide derived from human SMAD2 around the phosphorylation site of Ser465/467 CS(p-S)M(p-S)
• Antibody Type: Primary Antibody
• Concentration: 1mg/ml
• Dilution range: WB=1:500-2000, IHC-P=1:100-500, IHC-F=1:100-500, ICC/IF=1:100, IF=1:100-500, Flow-Cyt=1ug/Test
• Form/Appearance: Liquid
• Conjugation: Unconjugated
• MW: 58 kDa
• Target: SMAD2
• UniProt ID: Q15796
• RRID: AB_10929127
• Storage: Maintain refrigerated at 2-8°C for up to 2 weeks. For long term storage store at -20°C in small aliquots to prevent freeze-thaw cycles.
• Buffer/Preservatives: 0.01M TBS (pH7.4) with 1% rAlbumin, 0.02% Proclin300 and 50% Glycerol.
• Alternative names: Smad2(phospho S465+S467); phospho-Smad2(p-Ser465/4
• Note: For research use only
• Expiration Date: 12 months from date of receipt.

References

Zhang, Hongjun et al. Adenovirus‑mediated knockdown of activin A receptor type 2A attenuates immune‑induced hepatic fibrosis in mice and inhibits interleukin‑17‑induced activation of primary hepatic stellate cells Int J Mol Med, 42, 279-289 (2018)

PubMed: 29620144

Blank control (black line): Hela. Primary Antibody (green line): Rabbit Anti-Phospho-Smad2 (Ser465 + Ser467) antibody (orb6979), dilution: 1 ug/Test, Secondary Antibody (white blue line): Goat anti-rabbit IgG-AF488, dilution: 0.5 ug/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°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. Acquisition of 20000 events was performed.

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-Smad2 (Ser465 + Ser467)) polyclonal Antibody, Unconjugated (orb6979) 1:100, 90 minutes at 37°C, followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue) was used to stain the cell nuclei.

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-Smad2 (Ser465 + Ser467)) Polyclonal Antibody, Unconjugated (orb6979) at 1:400 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (Mouse testis), 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-Smad2 (Ser465 + Ser467)) Polyclonal Antibody, Unconjugated (orb6979) at 1:400 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructionsand DAB staining.

Sample: Lane 1: Placenta (Mouse) Lysate at 40 ug, Lane 2: Raw264.7 (Mouse) Cell Lysate at 30 ug, Lane 3: Testis (Rat) Lysate at 40 ug, Lane 4: Hela (Human) Cell Lysate at 30 ug, Lane 5: HT1080 (Human) Cell Lysate at 30 ug, Lane 6: Jurkat (Human) Cell Lysate at 30 ug, Lane 7: HL60 (Human) Cell Lysate at 30 ug, Primary: Anti-Phospho-Smad2 (Ser465 + Ser467) (orb6979) at 1/1000 dilution, Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution, Predicted band size: 60 kD, Observed band size: 60 kD.