HIF Prolyl Hydroxylases Recombinant Rabbit Monoclonal Antibody [JE63-25]
cat.: HA722468
| Product Type: | Recombinant Rabbit monoclonal IgG, primary antibodies |
|---|---|
| Species reactivity: | Human |
| Applications: | WB, IHC-P, IF-Cell, IP, FC |
| Clonality: | Monoclonal |
| Clone number: | JE63-25 |
| 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: 47 kDa |
| Isotype: | IgG |
| Immunogen: | Synthetic peptide within Human HIF Prolyl Hydroxylases aa 51-150 / 502. |
| Positive control: | HEK-293 cell lysate, A549 cell lysate, HCT 116 cell lysate, HeLa cell lysate, HEK-293, human lung cancer tissue, human colon cancer tissue. |
| Subcellular location: | Endoplasmic reticulum membrane. |
| Recommended Dilutions:
WB IHC-P IF-Cell IP FC |
1:1,000 1:200 1:100 1-2μg/sample 1:1,000 |
| Uniprot #: | SwissProt: Q9NXG6 Human |
| Alternative names: | EGLN4 FLJ20262 HIF prolyl hydroxylase PH4 HIF-PH4 HIF-prolyl hydroxylase 4 HIFPH4 HPH-4 Hypoxia inducible factor prolyl 4 hydroxylase Hypoxia inducible factor prolyl hydroxylase Hypoxia-inducible factor prolyl hydroxylase 4 P4H TM P4H with transmembrane domain P4H-TM P4htm P4HTM_HUMAN PH 4 PH4 PHD4 Proline 4 hydroxylase Prolyl 4 hydroxylase transmembrane (endoplasmic reticulum) Prolyl hydroxlase domain containing 4 Transmembrane prolyl 4-hydroxylase |
Images
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Fig1:
Western blot analysis of HIF Prolyl Hydroxylases on different lysates with Rabbit anti-HIF Prolyl Hydroxylases antibody (HA722468) at 1/1,000 dilution. Lane 1: HEK-293 cell lysate Lane 2: A549 cell lysate Lane 3: HCT 116 cell lysate Lane 4: HeLa cell lysate Lysates/proteins at 20 µg/Lane. Predicted band size: 47 kDa Observed band size: 47 kDa Exposure time: 25 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 (HA722468) 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:
Immunocytochemistry analysis of HEK-293 cells labeling HIF Prolyl Hydroxylases with Rabbit anti-HIF Prolyl Hydroxylases antibody (HA722468) 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-HIF Prolyl Hydroxylases antibody (HA722468) 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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Fig3:
Immunohistochemical analysis of paraffin-embedded human lung cancer tissue with Rabbit anti-HIF Prolyl Hydroxylases antibody (HA722468) at 1/1,000 dilution. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 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 (HA722468) 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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Fig4:
Immunohistochemical analysis of paraffin-embedded human colon cancer tissue with Rabbit anti-HIF Prolyl Hydroxylases antibody (HA722468) at 1/1,000 dilution. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 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 (HA722468) 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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Fig5:
HIF Prolyl Hydroxylases was immunoprecipitated from 0.2 mg HEK-293 cell lysate with HA722468 at 2 µg/25 µl agarose. Western blot was performed from the immunoprecipitate using HA722468 at 1/1,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature. Lane 1: HEK-293 cell lysate (input) Lane 2: HA722468 IP in HEK-293 cell lysate Lane 3: Rabbit IgG instead of HA722468 in HEK-293 cell lysate Blocking/Dilution buffer: 5% NFDM/TBST Exposure time: 26 seconds; ECL: K1801 |
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Fig6:
Flow cytometric analysis of HEK-293 cells labeling HIF Prolyl Hydroxylases. Cells were fixed and permeabilized. Then stained with the primary antibody (HA722468, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black). |
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