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Validation Data Gallery

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  • Western Blot (WB) analysis of various lysates using Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) Reco (80427-2-RR)

    WB analysis using 80427-2-RR (same clone as 80427-2-PBS)

    Non-treated and Calyculin A treated HEK-293 cells were subjected to SDS PAGE followed by western blot with 80427-2-RR (Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) antibody) at dilution of 1:2000 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with Alpha Tubulin (66031-1-Ig) antibody as a loading control. This data was developed using the same antibody clone with 80427-2-PBS in a different storage buffer formulation.

  • Immunofluorescence (IF) / fluorescent staining of HEK-293 cells using Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) Reco (80427-2-RR)

    IF Staining of HEK-293 using 80427-2-RR (same clone as 80427-2-PBS)

    Immunofluorescent analysis of (4% PFA) fixed TGF beta 1 treated and untreated HEK-293 cells using Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) antibody (80427-2-RR, Clone: 240826D11 ) at dilution of 1:500 and CoraLite®488-Conjugated Goat Anti-Rabbit IgG(H+L) (SA00013-2). This data was developed using the same antibody clone with 80427-2-PBS in a different storage buffer formulation.

  • Flow cytometry (FC) experiment of HEK-293 cells using Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) Reco (80427-2-RR)

    FC experiment of HEK-293 using 80427-2-RR (same clone as 80427-2-PBS)

    1X10^6 HEK-293 cells untreated (dashed lines) or treated with Calyculin A were intracellularly stained with 0.13 ug Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) Recombinant antibody (80427-2-RR, Clone:240826D11) and CoraLite®488-Conjugated Goat Anti-Rabbit IgG(H+L) (SA00013-2)(red), or 0.13 ug Rabbit IgG Isotype Control Recombinant Antibody (98136-1-RR, Clone: 240953C9) (blue). Cells were fixed with 4% PFA and permeabilized with 90% MeOH. This data was developed using the same antibody clone with 80427-2-PBS in a different storage buffer formulation.

  • ChIP experiment of HaCaT cells using Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) Reco (80427-2-RR)

    ChIP experiment of HaCaT using 80427-2-RR (same clone as 80427-2-PBS)

    Chromatin was prepared from HaCaT cells treated with TGF-β1 (7 ng/ml) for 1 h. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 20 µg of cross-linked chromatin, 5 µg of Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) (80427-2-RR) or 5 ug of Normal Rabbit IgG (98136-1-RR), and 20 µl of Protein A Magarose Beads. The immunoprecipitated DNA was quantified by real-time PCR. This data was developed using the same antibody clone with 80427-2-PBS in a different storage buffer formulation.

Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) Recombinant monoclonal antibody, PBS Only

Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) Uni-rAb® Recombinant Antibody for WB, IF/ICC, FC (Intra), ELISA, ChIP-qPCR
Cat No. 80427-2-PBS
Clone No.240826D11

Host / Isotype

Rabbit / IgG

Reactivity

human

Applications

WB, IF/ICC, FC (Intra), ELISA, ChIP-qPCR

Synonyms

P SMAD2, p SMAD2 (Ser465/467), P SMAD2 (Ser465/467)/SMAD3 (Ser423/425), P SMAD2 Ser465/467 /SMAD3 Ser423/425, p SMAD2/SMAD3

Formulation:  PBS Only
PBS and Azide
PBS Only
Conjugate:  Unconjugated
Size/Concentration: 

-/ -

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Tested Applications

Recommended dilution

ApplicationDilution
It is recommended that this reagent should be titrated in each testing system to obtain optimal results.

Product Information

80427-2-PBS targets Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) in WB, IF/ICC, FC (Intra), ELISA, ChIP-qPCR applications and shows reactivity with human samples.

Tested Reactivity human
Host / Isotype Rabbit / IgG
Class Recombinant
Type Antibody
Immunogen

Peptide

相同性解析による交差性が予測される生物種
Full Name SMAD family member 2
Calculated molecular weight 467 aa, 52 kDa
Observed molecular weight60 kDa
GenBank accession numberBC014840
Gene Symbol SMAD2
Gene ID (NCBI) 4087
Conjugate Unconjugated
Form
FormLiquid
Purification MethodProtein A purfication
UNIPROT IDQ15796
Storage Buffer PBS only{{ptg:BufferTemp}}7.3
Storage ConditionsStore at -80°C.

Background Information

SMAD2, also named as MADH2 and MADR2, belongs to the dwarfin/SMAD family, contains 1 MH1 (MAD homology 1) domain and 1 MH2 (MAD homology 2) domain. SMAD2 is a receptor-regulated SMAD(R-SMAD) that is an intracellular signal transducer and transcriptional modulator activated by TGF-beta and activin type 1 receptor kinases. This protein may act as a tumor suppressor in colorectal carcinoma. It is phosphorylated on one or several of Thr-220, Ser-245, Ser-250, and Ser-255. In response to TGF-beta, It is phosphorylated on Ser-465/467 by TGF-beta and activin type 1 receptor kinases, and then able to interact with SMURF2, recruiting other proteins, such as SNON, for degradation. In response to decorin, the naturally occurring inhibitor of TGF-beta signaling, it is phosphorylated on Ser-240 by CaMK2. It is phosphorylated by MAPK3 upon EGF stimulation; which increases transcriptional activity and stability, and is blocked by calmodulin. In response to TGF-beta, it is ubiquitinated by NEDD4L, which promotes its degradation. In response to TGF-beta signaling, it is acetylated on Lys-19 by coactivators, which increases transcriptional activity. The molecular weight of unphosphorylated forms of Smad2 is 52 kDa and phosphorylated forms of Smad2 is 58 kDa. (PMID: 9006934)