Phospho-Smad1/5/9 (S463/S465/S467) Recombinant Rabbit Monoclonal Antibody [JE59-46]
cat.: HA722566
| Product Type: | Recombinant Rabbit monoclonal IgG, primary antibodies |
|---|---|
| Species reactivity: | Human, Mouse, Rat |
| Applications: | WB, IHC, IF-Cell |
| Clonality: | Monoclonal |
| Clone number: | JE59-46 |
| Form: | Liquid |
| Storage condition: | Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles. |
| Storage buffer: | 1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide. |
| Concentration: | 1ug/ul |
| Purification: | Protein A affinity purified. |
| Molecular weight: | Predicted band size: 52 kDa |
| Isotype: | IgG |
| Immunogen: | Synthetic phosphopeptide corresponding to residues surrounding Ser463/465 of human SMAD1 and SMAD5 protein. |
| Positive control: | HeLa treated with 10ng/mL BMP4 for 1 hour cell lysate, NIH/3T3 treated with 10ng/mL BMP4 for 1 hour cell lysate, human testis tissue, C6 treated with 10ng/mL BMP4 for 1 hour cell lysate, HeLa cells treated with 10ng/mL BMP4 for 1 hour, NIH/3T3 cells treated with 10ng/mL BMP4 for 1 hour. |
| Subcellular location: | Cytoplasm, Nucleus, Mitochondrion. |
| Recommended Dilutions:
WB IHC IF-Cell |
1:1,000 1:1,000 1:100 |
| Uniprot #: | SwissProt: Q15797 Human | Q99717 Human | O15198 Human | P70340 Mouse | P97454 Mouse | Q9JIW5 Mouse | O54835 Rat | P97588 Rat | Q9R1V3 Rat |
| Alternative names: | BSP-1 BSP1 HsMAD1 JV4-1 JV41 MAD homolog 1 MAD mothers against decapentaplegic homolog 1 Mad related protein 1 Mad-related protein 1 MADH1 MADR1 Mothers against decapentaplegic homolog 1 Mothers against DPP homolog 1 SMA- AND MAD-RELATED PROTEIN 1 SMAD 1 SMAD family member 1 SMAD mothers against DPP homolog 1 Smad1 SMAD1_HUMAN TGF beta signaling protein 1 Transforming growth factor-beta-signaling protein 1 DKFZp781C1895 DKFZp781O1323 Dwfc hSmad5 JV5 1 JV5-1 MAD homolog 5 MAD, mothers against decapentaplegic homolog 5 MADH 5 MADH5 Mothers against decapentaplegic homolog 5 mothers against decapentaplegic, drosophila, homolog of, 5 Mothers against DPP homolog 5 MusMLP SMA and MAD related protein 5 SMAD 5 SMAD family member 5 SMAD, mothers against DPP homolog 5 Smad5 SMAD5_HUMAN MAD homolog 9 Madh6 Mothers against decapentaplegic Mothers against decapentaplegic homolog 9 Mothers against DPP homolog 9 ...... |
Images
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Fig1:
Western blot analysis of Phospho-Smad1/5/9 (S463/S465/S467) on different lysates with Rabbit anti-Phospho-Smad1/5/9 (S463/S465/S467) antibody (HA722566) at 1/1,000 dilution. Lane 1: HeLa cell lysate Lane 2: HeLa treated with 10ng/mL BMP4 for 1 hour cell lysate Lane 3: HeLa treated with 10ng/mL BMP4 for 1 hour cell lysate, then the membrane treated with λpp for 1 hour Lane 4: NIH/3T3 cell lysate Lane 5: NIH/3T3 treated with 10ng/mL BMP4 for 1 hour cell lysate Lysates/proteins at 20 µg/Lane. Predicted band size: 52 kDa Observed band size: 52 kDa Exposure time: Lane 1-3: 3 minutes; Lane 4-5: 42 seconds; ECL: K1801; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722566) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. |
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Fig2:
Immunohistochemical analysis of paraffin-embedded human testis tissue with Rabbit anti-Phospho-Smad1/5/9 (S463/S465/S467) antibody (HA722566) at 1/1,000 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA722566) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. |
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Fig3:
Western blot analysis of Phospho-Smad1/5/9 (S463/S465/S467) on different lysates with Rabbit anti-Phospho-Smad1/5/9 (S463/S465/S467) antibody (HA722566) at 1/1,000 dilution. Lane 1: C6 cell lysate Lane 2: C6 treated with 10ng/mL BMP4 for 1 hour cell lysate Lane 3: C6 treated with 10ng/mL BMP4 for 1 hour cell lysate, then the membrane treated with λpp for 1 hour Lysates/proteins at 10 µg/Lane. Predicted band size: 52 kDa Observed band size: 52 kDa Exposure time: 1 minute 18 seconds; ECL: K1801; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722566) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. |
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Fig4:
Immunocytochemistry analysis of HeLa cells treated with 10ng/mL BMP4 for 1 hour labeling Phospho-Smad1/5/9 (S463/S465/S467) with Rabbit anti-Phospho-Smad1/5/9 (S463/S465/S467) antibody (HA722566) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-Smad1/5/9 (S463/S465/S467) antibody (HA722566) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. |
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Fig5:
Immunocytochemistry analysis of NIH/3T3 cells treated with 10ng/mL BMP4 for 1 hour labeling Phospho-Smad1/5/9 (S463/S465/S467) with Rabbit anti-Phospho-Smad1/5/9 (S463/S465/S467) antibody (HA722566) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-Smad1/5/9 (S463/S465/S467) antibody (HA722566) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. |
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