Phospho-PKA R2 (S99) Recombinant Rabbit Monoclonal Antibody [SC54-04]
cat.: ET1610-29
| Product Type: | Recombinant Rabbit monoclonal IgG, primary antibodies |
|---|---|
| Species reactivity: | Human, Mouse, Rat, Pig |
| Applications: | WB, IF-Cell, IF-Tissue, IHC-P, IP, FC |
| Clonality: | Monoclonal |
| Clone number: | SC54-04 |
| Form: | Liquid |
| Storage condition: | Store at +4℃ after thawing. Aliquot store at -20℃ or -80℃. 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: 46 kDa |
| Isotype: | IgG |
| Immunogen: | Synthetic phospho-peptide corresponding to residues surrounding Ser99 of Human PKA R2 aa 82-126 / 404. |
| Positive control: | PC-12 treated with 100nM Calyculin A for 30 minutes cell lysate, human kidney tissue, mouse kidney tissue, rat kidney tissue, human colon cancer tissue, PC-12 cells treated with 100nM Calyculin A for 30 minutes, PC-12. |
| Subcellular location: | Cytoplasm, Cell membrane. |
| Recommended Dilutions:
WB IF-Cell IF-Tissue IHC-P IP FC |
1:2,000 1:200 1:50-1:200 1:1,000 1-2μg/sample 1:1,000 |
| Uniprot #: | SwissProt: P13861 Human | P12367 Mouse | P12368 Rat |
| Alternative names: | cAMP dependent protein kinase regulatory subunit alpha 2 cAMP dependent protein kinase regulatory subunit RII alpha cAMP dependent protein kinase type II alpha regulatory chain cAMP dependent protein kinase type II alpha regulatory subunit cAMP-dependent protein kinase type II-alpha regulatory subunit KAP2 KAP2_HUMAN MGC3606 PKR 2 PKR2 PRKA R2 PRKAR 2 PRKAR2 PRKAR2A Protein kinase A RII alpha subunit Protein kinase cAMP dependent regulatory type II alpha |
Images
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Fig1:
Western blot analysis of Phospho-PKA R2 (S99) on different lysates with Rabbit anti-Phospho-PKA R2 (S99) antibody (ET1610-29) at 1/2,000 dilution. Lane 1: PC-12 cell lysate Lane 2: PC-12 treated with 100nM Calyculin A for 30 minutes cell lysate Lysates/proteins at 20 µg/Lane. Predicted band size: 46 kDa Observed band size: 50 kDa Exposure time: 20 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 (ET1610-29) at 1/2,000 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 kidney tissue untreated / treated with λpp with Rabbit anti-Phospho-PKA R2 (S99) antibody (ET1610-29) 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 (ET1610-29) 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:
Immunohistochemical analysis of paraffin-embedded mouse kidney tissue untreated / treated with λpp with Rabbit anti-Phospho-PKA R2 (S99) antibody (ET1610-29) 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 (ET1610-29) 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 rat kidney tissue untreated / treated with λpp with Rabbit anti-Phospho-PKA R2 (S99) antibody (ET1610-29) 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 (ET1610-29) 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:
Immunohistochemical analysis of paraffin-embedded human colon cancer tissue with Rabbit anti-Phospho-PKA R2 (S99) antibody (ET1610-29) 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 (ET1610-29) 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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Fig6:
Immunocytochemistry analysis of PC-12 cells untreated / treated with 100nM Calyculin A for 30 minutes labeling Phospho-PKA R2 (S99) with Rabbit anti-Phospho-PKA R2 (S99) antibody (ET1610-29) at 1/200 dilution. Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 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-PKA R2 (S99) antibody (ET1610-29) at 1/200 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 (HA601187, 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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Fig7:
Flow cytometric analysis of PC-12 cells labeling Phospho-PKA R2 (S99). Cells were fixed and permeabilized. Then stained with the primary antibody (ET1610-29, 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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Fig8:
Phospho-PKA R2 (S99) was immunoprecipitated from 0.2 mg PC-12 cell lysate with ET1610-29 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using ET1610-29 at 1/1,000 dilution. HRP Conjugated Anti-Rabbit IgG for IP secondary antibody at 1/5,000 dilution was used for 1 hour at room temperature. Lane 1: PC-12 cell lysate (input) Lane 2: ET1610-29 IP in PC-12 cell lysate Lane 3: Rabbit IgG instead of ET1610-29 in PC-12 cell lysate Blocking/Dilution buffer: 5% NFDM/TBST Exposure time: 20 seconds; ECL: K1801 |
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.