Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD3 (pS423/425) + SMAD5 (pS463/465 ) + SMAD1 (pS463/465) + SMAD2 (pS465/467) antibody [EP823Y] - BSA and Azide free (AB172202)

Immunohistochemical analysis of Smad3 in paraffin embedded human liver carcinoma tissue using ab52903 at 1/100 dilution.This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52903).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

Immunocytochemistry/Immunofluorescence analysis of A549 +/- TGFβ (5ng/ml 24h) and A549 + TGFβ (5ng/ml 24h) + Lamda phosphatase (LP) cells. Smad3 (phospho S423 + S425) was labelled with purified ab52903 at a dilution of 1/100 dilution while Smad3 was labelled with ab207447 at a dilution of 1/500 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% triton X-100. ab150077 (goat anti-rabbit IgG Alexa Fluor^® 488) (1/1000 dilution) was used as the secondary antibody. The cells were co-stained with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution. Nuclei counterstained with DAPI (blue). Control : PBS instead of the primary antibody.

This ICC/IF data was generated using the same anti-phospho Smad3 S423/425 antibody clone, EP823Y, in a different buffer formulation (ab52903).

• ICC/IF

AbReview40862****

Immunocytochemistry/ Immunofluorescence - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

ab52903 staining Smad3 in mouse primary embryonic epicardial cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% formaldehyde permeabilized with 0.5% Triton X-100 and blocked with PBS + 1% BSA + 10% goat serum + 0.1% Triton X-100 for 1 hour at 20°C. Samples were incubated with primary antibody (1/100 dilution in PBS + 1% BSA + 10% goat serum + 0.1% Triton X-100) for 16 hours at 4°C. An Alexa Fluor^®488-conjugated goat anti-rabbit IgG polyclonal (1/200 dilution) was used as the secondary antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52903).

This image is courtesy of an anonymous Abreview.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD3 (pS423/425) + SMAD5 (pS463/465 ) + SMAD1 (pS463/465) + SMAD2 (pS465/467) antibody [EP823Y] - BSA and Azide free (AB172202)

This IHC data was generated using the same anti-phospho Smad3 S423/425 antibody clone, EP823Y, in a different buffer formulation (cat# ab52903).

ab52903 staining Smad3 in Human stomach tissue sections by Immunohistochemistry (Formalin/PFA fixed paraffin embedded sections). Tissue was fixed with paraffin and antigen retrieval was by heat mediation using ab93684 (Tris/EDTA buffer, Ph9.0). Samples were incubated with primary antibody at a 1/200 dilution. A ready to use rabbit specific IHC polymer detection kit HRP/DAP (ab209101). Hematoxylin was used as a counterstain. Nuclear and weakly cytoplasmic staining on human stomach without alkaline phosphatase treatment (image A). No signal can be detected when tissues were treated with alkaline phosphatase (image B).

• ICC/IF

AbReview16498****

Immunocytochemistry/ Immunofluorescence - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

ab52903 staining Smad3 (phospho S423 + S425) in human TII Pneumocyte A549 cells by Immunocytochemistry/ Immunofluorescence. Cells were fixed with paraformaldehyde and permeabilized with 0.1% Triton x100 before blocking with 3% BSA for 1 hour at RT. Samples were incubated with primary antibody (1/200 : in 3% BSA in 1x PBST) for 24 hours at 4°C. A TRITC-conjugated goat polyclonal to rabbit IgG was used as secondary antibody at 1/200 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52903).

This image is courtesy of an Abreview submitted by Aaron Gardner.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD3 (pS423/425) + SMAD5 (pS463/465 ) + SMAD1 (pS463/465) + SMAD2 (pS465/467) antibody [EP823Y] - BSA and Azide free (AB172202)

Purified ab52903 staining Smad3 in Mouse kidney tissue sections by Immunohistochemistry (Formalin/PFA fixed paraffin embedded sections). Tissue was fixed with paraffin and antigen retrieval was by heat mediation using ab93684 (Tris/EDTA buffer, pH 9.0). Samples were incubated with primary antibody at a 1/200 dilution. A ready to use rabbit specific IHC polymer detection kit HRP/DAP (ab209101). Hematoxylin was used as a counterstain. Nuclear and weakly cytoplasmic staining on mouse kidney without alkaline phosphatase treatment (image A). No signal can be detected when tissues were treated with alkaline phosphatase (image B).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52903).

• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

TGF-β1 signaling is impaired in NDRG1-silenced MEFs. PML^+/+ mouse embryonic fibroblasts (MEFs) were transfected with either CTL-siRNAs (A & B) or NDRG1-siRNAs (C & D) and induced with100 ng/ml TGF-β1. Immunofluorescent staining revealed intense nuclear staining for phosphorylated SMAD3 (SMAD3-P ab52903) in CTL-siRNA treated MEFs (B) while only weak nuclear staining for MEFs treated with NDRG1-siRNA (D).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52903).

Image from Tang MK et al. PLoS One. 2013;8(3):e59477. Fig 12.; doi: 10.1371/journal.pone.0059477.

• Dot

Unknown

Dot Blot - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

Dot blot analysis of Smad 3 (phospho S423 + S425) phospho peptide (Lane 1), Smad 3 (phospho S423) phospho peptide (Lane 2), Smad 3 (phospho S425) phospho peptide (Lane 3) and Smad 3 non-phospho peptide (Lane 4) labelling Smad 3 (phospho S423 + S425) with ab52903 at a dilution of 1/1000. A Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) was used as the secondary antibody at a dilution of 1/20,000. Blocking and dilution buffer : 5% NFDM /TBST.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52903).

• ChIC/CUT&RUN-seq

Lab

ChIC/CUT&RUN sequencing - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

This data was developed using the same antibody clone in a different buffer formulation (ab52903). ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 A549 (Human lung carcinoma cell line) cells treated with hTGF-β1 (7 ng/mL 1 h) and 5 µg of ab52903 [EP823Y]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown. The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

• ChIC/CUT&RUN-seq

Lab

ChIC/CUT&RUN sequencing - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

This data was developed using the same antibody clone in a different buffer formulation (ab52903). ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 A549 (Human lung carcinoma cell line) cells treated with hTGF-β1 (7 ng/mL 1 h) and 5 µg of ab52903 [EP823Y]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown. The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

• ChIC/CUT&RUN-seq

Lab

ChIC/CUT&RUN sequencing - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

This data was developed using the same antibody clone in a different buffer formulation (ab52903). ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 A549 (Human lung carcinoma cell line) cells treated with hTGF-β1 (7 ng/mL 1 h) and 5 µg of ab52903 [EP823Y]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown. The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

• IHC-P

PubMed

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD3 (pS423/425) + SMAD5 (pS463/465 ) + SMAD1 (pS463/465) + SMAD2 (pS465/467) antibody [EP823Y] - BSA and Azide free (AB172202)

Representative IHC photomicrographs from an Environmental enteropathy (EE) duodenal biopsy showing p-SMAD3 staining (ab52903) in only the epithelium (arrows).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab52903).

Image from Syed S et al. PLoS Negl Trop Dis. 2018;12(2):e0006224. Fig 4.; doi: 10.1371/journal.pntd.0006224.

• Dot

Lab

Dot Blot - Anti-SMAD3 (pS423/425)+SMAD5&SMAD1 (pS463/465)+SMAD2 (pS465/467) antibody [EP823Y] - BSA&Azide free (AB172202)

Blocking and diluting buffer and concentration : 5% NFDM /TBST

All lanes:

Dot Blot - Anti-SMAD3 (pS423/425) + SMAD5 (pS463/465) + SMAD1 (pS463/465) + SMAD2 (pS465/467) antibody [EP823Y] (<a href='/en-us/products/primary-antibodies/smad3-ps423-425-smad5-ps463-465-smad1-ps463-465-smad2-ps465-467-antibody-ep823y-ab52903'>ab52903</a>) at 1/1000 dilution

Lane 1:

Smad5 non-phospho peptide

Lane 2:

Smad5 (s463+s465) phospho peptide

Lane 3:

Smad2 non-phospho peptide

Lane 4:

Smad2 (s465+s467) phospho peptide

Lane 5:

Smad1 non-phospho peptide

Lane 6:

Smad1 (s463+s465) phospho peptide

Lane 7:

Smad3 non-phospho peptide

Lane 8:

Smad3 (s423+s435) phospho peptide

Secondary

All lanes:

Dot Blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

false

Exposure time: 59s