![Immunocytochemistry/Immunofluorescence: NT5C Antibody [NBP1-84563] - Staining of human cell line U-2 OS shows localization to cytosol. Antibody staining is shown in green.](http://images.novusbio.com/fullsize/NT5C-Antibody-Immunocytochemistry-Immunofluorescence-NBP1-84563-img0011.jpg "Immunocytochemistry/Immunofluorescence: NT5C Antibody [NBP1-84563] - Staining of human cell line U-2 OS shows localization to cytosol. Antibody staining is shown in green.")
| Reactivity | HuSpecies Glossary |
| Applications | WB, Simple Western, ICC/IF, IHC, Mycoplasma |
| Clonality | Polyclonal |
| Host | Rabbit |
| Conjugate | Unconjugated |
| Format | BSA Free |
| Description | Novus Biologicals Rabbit NT5C Antibody - BSA Free (NBP1-84563) is a polyclonal antibody validated for use in IHC, WB, ICC/IF and Simple Western. Anti-NT5C Antibody: Cited in 1 publication. All Novus Biologicals antibodies are covered by our 100% guarantee. |
| Immunogen | This antibody was developed against Recombinant Protein corresponding to amino acids: VLADFEAGLLRGFRRRFPEEPHVPLEQRRGFLAREQYRALRPDLADKVASVYEAPGFFLDLEPIP |
| Isotype | IgG |
| Clonality | Polyclonal |
| Host | Rabbit |
| Gene | NT5C |
| Purity | Affinity purified |
| Innovator's Reward | Test in a species/application not listed above to receive a full credit towards a future purchase. |
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| Application Notes | ICC/IF Fixation Permeabilization: Use PFA/Triton X-100. IHC-Paraffin HIER pH6 retrieval is recommended. See Simple Western Antibody Database for Simple Western validation: Tested in U-2OS, RT-4, separated by Size, antibody dilution of 1:20, apparent MW was 29 kDa |
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| Publications |
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| Storage | Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles. |
| Buffer | PBS (pH 7.2) and 40% Glycerol |
| Preservative | 0.02% Sodium Azide |
| Purity | Affinity purified |
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| Publication using NBP1-84563 | Applications | Species |
|---|---|---|
| Moniz LS, Surinova S, Ghazaly E et al. Phosphoproteomic comparison of Pik3ca and Pten signalling identifies the nucleotidase NT5C as a novel AKT substrate Sci Rep 2017-01-06 [PMID: 28059163] (WB, Human) | WB | Human |
Secondary Antibodies |
Isotype Controls |
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| Gene Symbol | NT5C |
![Simple Western: NT5C Antibody [NBP1-84563] - Simple Western lane view shows a specific band for NT5C in 0.2 mg/ml of RT-4 lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.](http://images.novusbio.com/fullsize/NT5C-Antibody-Simple-Western-NBP1-84563-img0007.jpg "Simple Western: NT5C Antibody [NBP1-84563] - Simple Western lane view shows a specific band for NT5C in 0.2 mg/ml of RT-4 lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.")
![Simple Western: NT5C Antibody [NBP1-84563] - Simple Western lane view shows a specific band for NT5C in 0.2 mg/ml of U-2OS lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.](http://images.novusbio.com/fullsize/NT5C-Antibody-Simple-Western-NBP1-84563-img0008.jpg "Simple Western: NT5C Antibody [NBP1-84563] - Simple Western lane view shows a specific band for NT5C in 0.2 mg/ml of U-2OS lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.")
![Simple Western: NT5C Antibody [NBP1-84563] - Electropherogram image(s) of corresponding Simple Western lane view. NT5C antibody was used at 1:20 dilution on U-2OS lysates(s).](http://images.novusbio.com/fullsize/NT5C-Antibody-Simple-Western-NBP1-84563-img0009.jpg "Simple Western: NT5C Antibody [NBP1-84563] - Electropherogram image(s) of corresponding Simple Western lane view. NT5C antibody was used at 1:20 dilution on U-2OS lysates(s).")
![Western Blot: NT5C Antibody [NBP1-84563] - Regulation of NT5C S184 phosphorylation.(a) Sensitivity of S184 phosphorylation to PI3K pathway inhibitors. Primary MEFs were treated with A66 (3 μM), MK-2206 (1 μM), Rapamycin (20 nM), U0126 (10 μM) or DMSO as control for 3 h or overnight (o/n) for 16–18 h before lysis. (left) Lysates were immunoblotted as indicated. (right) Quantification of 4 independent experiments. Values are expressed relative to DMSO treated Pik3caH1047R/WT cells. Error bars are sem, **p < 0.01. (b) Sensitivity of S184 phosphorylation to growth factor deprivation. Indicated primary MEFs were grown in 10% FBS or starved overnight in 0.1% FBS. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 5 independent experiments. Values are expressed relative to Pik3caH1047R/WT cells grown in 10% FBS. Error bars are sem, ***p < 0.001. (c) Sensitivity of S184 phosphorylation to growth factor stimulation. Primary MEFs were starved overnight in 0.1% FBS & stimulated with FBS (10%), Insulin (100 nM) or EGF (100 ng/ml) for the indicated times before lysis. DMSO or MK-2206 (1 μM) was added at same time as stimuli. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 3–5 independent experiments. Values are expressed relative to time 0 for each stimuli. Error bars are sem. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.](http://images.novusbio.com/fullsize/nbp1-84563_rabbit-polyclonal-nt5c-antibody-31020241612237.jpg "Western Blot: NT5C Antibody [NBP1-84563] - Regulation of NT5C S184 phosphorylation.(a) Sensitivity of S184 phosphorylation to PI3K pathway inhibitors. Primary MEFs were treated with A66 (3 μM), MK-2206 (1 μM), Rapamycin (20 nM), U0126 (10 μM) or DMSO as control for 3 h or overnight (o/n) for 16–18 h before lysis. (left) Lysates were immunoblotted as indicated. (right) Quantification of 4 independent experiments. Values are expressed relative to DMSO treated Pik3caH1047R/WT cells. Error bars are sem, **p < 0.01. (b) Sensitivity of S184 phosphorylation to growth factor deprivation. Indicated primary MEFs were grown in 10% FBS or starved overnight in 0.1% FBS. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 5 independent experiments. Values are expressed relative to Pik3caH1047R/WT cells grown in 10% FBS. Error bars are sem, ***p < 0.001. (c) Sensitivity of S184 phosphorylation to growth factor stimulation. Primary MEFs were starved overnight in 0.1% FBS & stimulated with FBS (10%), Insulin (100 nM) or EGF (100 ng/ml) for the indicated times before lysis. DMSO or MK-2206 (1 μM) was added at same time as stimuli. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 3–5 independent experiments. Values are expressed relative to time 0 for each stimuli. Error bars are sem. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.")
![Western Blot: NT5C Antibody [NBP1-84563] - Impact of S184 phosphorylation of NT5C catalytic activity.(a) S184 phosphorylation does not regulate NT5C nucleotidase activity in vitro. (left) Immunoprecipitates of Flag-NT5C ectopically expressed in HEK293 cells were incubated with 5 mM of the indicated nucleotides. Phosphate release was measured using a malachite green colorimetric assay & expressed as a percent of total nucleotide. The experiment was performed in duplicate & repeated 3 times independently. Error bars are sem. (right) Representative immunoblot from experimental cells. (b) Cells expressing Pik3caH1047R have elevated dNTP levels. (left, middle) Nucleotides were extracted from primary MEFs & analysed by UPLC-MS/MS. The experiment was performed in triplicate & repeated 4 times independently. Error bars are sem, *p < 0.05, **p < 0.01. (right) Representative immunoblot from experimental cells. (c) Effect of NT5C knockdown on cellular dCTP levels. (left) Nucleotides were extracted from primary MEFs, stably expressing indicated siRNA, & analysed by UPLC-MS/MS. The experiment was performed in triplicate & repeated 3 times independently. Error bars are sem. (right) Representative immunoblot from experimental cells. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.](http://images.novusbio.com/fullsize/nbp1-84563_rabbit-polyclonal-nt5c-antibody-310202416121989.jpg "Western Blot: NT5C Antibody [NBP1-84563] - Impact of S184 phosphorylation of NT5C catalytic activity.(a) S184 phosphorylation does not regulate NT5C nucleotidase activity in vitro. (left) Immunoprecipitates of Flag-NT5C ectopically expressed in HEK293 cells were incubated with 5 mM of the indicated nucleotides. Phosphate release was measured using a malachite green colorimetric assay & expressed as a percent of total nucleotide. The experiment was performed in duplicate & repeated 3 times independently. Error bars are sem. (right) Representative immunoblot from experimental cells. (b) Cells expressing Pik3caH1047R have elevated dNTP levels. (left, middle) Nucleotides were extracted from primary MEFs & analysed by UPLC-MS/MS. The experiment was performed in triplicate & repeated 4 times independently. Error bars are sem, *p < 0.05, **p < 0.01. (right) Representative immunoblot from experimental cells. (c) Effect of NT5C knockdown on cellular dCTP levels. (left) Nucleotides were extracted from primary MEFs, stably expressing indicated siRNA, & analysed by UPLC-MS/MS. The experiment was performed in triplicate & repeated 3 times independently. Error bars are sem. (right) Representative immunoblot from experimental cells. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.")
![Western Blot: NT5C Antibody [NBP1-84563] - Regulation of NT5C S184 phosphorylation.(a) Sensitivity of S184 phosphorylation to PI3K pathway inhibitors. Primary MEFs were treated with A66 (3 μM), MK-2206 (1 μM), Rapamycin (20 nM), U0126 (10 μM) or DMSO as control for 3 h or overnight (o/n) for 16–18 h before lysis. (left) Lysates were immunoblotted as indicated. (right) Quantification of 4 independent experiments. Values are expressed relative to DMSO treated Pik3caH1047R/WT cells. Error bars are sem, **p < 0.01. (b) Sensitivity of S184 phosphorylation to growth factor deprivation. Indicated primary MEFs were grown in 10% FBS or starved overnight in 0.1% FBS. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 5 independent experiments. Values are expressed relative to Pik3caH1047R/WT cells grown in 10% FBS. Error bars are sem, ***p < 0.001. (c) Sensitivity of S184 phosphorylation to growth factor stimulation. Primary MEFs were starved overnight in 0.1% FBS & stimulated with FBS (10%), Insulin (100 nM) or EGF (100 ng/ml) for the indicated times before lysis. DMSO or MK-2206 (1 μM) was added at same time as stimuli. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 3–5 independent experiments. Values are expressed relative to time 0 for each stimuli. Error bars are sem. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.](http://images.novusbio.com/fullsize/nbp1-84563_rabbit-polyclonal-nt5c-antibody-310202416113523.jpg "Western Blot: NT5C Antibody [NBP1-84563] - Regulation of NT5C S184 phosphorylation.(a) Sensitivity of S184 phosphorylation to PI3K pathway inhibitors. Primary MEFs were treated with A66 (3 μM), MK-2206 (1 μM), Rapamycin (20 nM), U0126 (10 μM) or DMSO as control for 3 h or overnight (o/n) for 16–18 h before lysis. (left) Lysates were immunoblotted as indicated. (right) Quantification of 4 independent experiments. Values are expressed relative to DMSO treated Pik3caH1047R/WT cells. Error bars are sem, **p < 0.01. (b) Sensitivity of S184 phosphorylation to growth factor deprivation. Indicated primary MEFs were grown in 10% FBS or starved overnight in 0.1% FBS. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 5 independent experiments. Values are expressed relative to Pik3caH1047R/WT cells grown in 10% FBS. Error bars are sem, ***p < 0.001. (c) Sensitivity of S184 phosphorylation to growth factor stimulation. Primary MEFs were starved overnight in 0.1% FBS & stimulated with FBS (10%), Insulin (100 nM) or EGF (100 ng/ml) for the indicated times before lysis. DMSO or MK-2206 (1 μM) was added at same time as stimuli. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 3–5 independent experiments. Values are expressed relative to time 0 for each stimuli. Error bars are sem. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.")
![Western Blot: NT5C Antibody [NBP1-84563] - Impact of S184 phosphorylation of NT5C catalytic activity.(a) S184 phosphorylation does not regulate NT5C nucleotidase activity in vitro. (left) Immunoprecipitates of Flag-NT5C ectopically expressed in HEK293 cells were incubated with 5 mM of the indicated nucleotides. Phosphate release was measured using a malachite green colorimetric assay & expressed as a percent of total nucleotide. The experiment was performed in duplicate & repeated 3 times independently. Error bars are sem. (right) Representative immunoblot from experimental cells. (b) Cells expressing Pik3caH1047R have elevated dNTP levels. (left, middle) Nucleotides were extracted from primary MEFs & analysed by UPLC-MS/MS. The experiment was performed in triplicate & repeated 4 times independently. Error bars are sem, *p < 0.05, **p < 0.01. (right) Representative immunoblot from experimental cells. (c) Effect of NT5C knockdown on cellular dCTP levels. (left) Nucleotides were extracted from primary MEFs, stably expressing indicated siRNA, & analysed by UPLC-MS/MS. The experiment was performed in triplicate & repeated 3 times independently. Error bars are sem. (right) Representative immunoblot from experimental cells. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.](http://images.novusbio.com/fullsize/nbp1-84563_rabbit-polyclonal-nt5c-antibody-31020241612211.jpg "Western Blot: NT5C Antibody [NBP1-84563] - Impact of S184 phosphorylation of NT5C catalytic activity.(a) S184 phosphorylation does not regulate NT5C nucleotidase activity in vitro. (left) Immunoprecipitates of Flag-NT5C ectopically expressed in HEK293 cells were incubated with 5 mM of the indicated nucleotides. Phosphate release was measured using a malachite green colorimetric assay & expressed as a percent of total nucleotide. The experiment was performed in duplicate & repeated 3 times independently. Error bars are sem. (right) Representative immunoblot from experimental cells. (b) Cells expressing Pik3caH1047R have elevated dNTP levels. (left, middle) Nucleotides were extracted from primary MEFs & analysed by UPLC-MS/MS. The experiment was performed in triplicate & repeated 4 times independently. Error bars are sem, *p < 0.05, **p < 0.01. (right) Representative immunoblot from experimental cells. (c) Effect of NT5C knockdown on cellular dCTP levels. (left) Nucleotides were extracted from primary MEFs, stably expressing indicated siRNA, & analysed by UPLC-MS/MS. The experiment was performed in triplicate & repeated 3 times independently. Error bars are sem. (right) Representative immunoblot from experimental cells. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.")
![Western Blot: NT5C Antibody [NBP1-84563] - Regulation of NT5C S184 phosphorylation.(a) Sensitivity of S184 phosphorylation to PI3K pathway inhibitors. Primary MEFs were treated with A66 (3 μM), MK-2206 (1 μM), Rapamycin (20 nM), U0126 (10 μM) or DMSO as control for 3 h or overnight (o/n) for 16–18 h before lysis. (left) Lysates were immunoblotted as indicated. (right) Quantification of 4 independent experiments. Values are expressed relative to DMSO treated Pik3caH1047R/WT cells. Error bars are sem, **p < 0.01. (b) Sensitivity of S184 phosphorylation to growth factor deprivation. Indicated primary MEFs were grown in 10% FBS or starved overnight in 0.1% FBS. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 5 independent experiments. Values are expressed relative to Pik3caH1047R/WT cells grown in 10% FBS. Error bars are sem, ***p < 0.001. (c) Sensitivity of S184 phosphorylation to growth factor stimulation. Primary MEFs were starved overnight in 0.1% FBS & stimulated with FBS (10%), Insulin (100 nM) or EGF (100 ng/ml) for the indicated times before lysis. DMSO or MK-2206 (1 μM) was added at same time as stimuli. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 3–5 independent experiments. Values are expressed relative to time 0 for each stimuli. Error bars are sem. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.](http://images.novusbio.com/fullsize/nbp1-84563_rabbit-polyclonal-nt5c-antibody-310202416114732.jpg "Western Blot: NT5C Antibody [NBP1-84563] - Regulation of NT5C S184 phosphorylation.(a) Sensitivity of S184 phosphorylation to PI3K pathway inhibitors. Primary MEFs were treated with A66 (3 μM), MK-2206 (1 μM), Rapamycin (20 nM), U0126 (10 μM) or DMSO as control for 3 h or overnight (o/n) for 16–18 h before lysis. (left) Lysates were immunoblotted as indicated. (right) Quantification of 4 independent experiments. Values are expressed relative to DMSO treated Pik3caH1047R/WT cells. Error bars are sem, **p < 0.01. (b) Sensitivity of S184 phosphorylation to growth factor deprivation. Indicated primary MEFs were grown in 10% FBS or starved overnight in 0.1% FBS. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 5 independent experiments. Values are expressed relative to Pik3caH1047R/WT cells grown in 10% FBS. Error bars are sem, ***p < 0.001. (c) Sensitivity of S184 phosphorylation to growth factor stimulation. Primary MEFs were starved overnight in 0.1% FBS & stimulated with FBS (10%), Insulin (100 nM) or EGF (100 ng/ml) for the indicated times before lysis. DMSO or MK-2206 (1 μM) was added at same time as stimuli. (top) Lysates were immunoblotted as indicated. (bottom) Quantification of 3–5 independent experiments. Values are expressed relative to time 0 for each stimuli. Error bars are sem. Image collected & cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.")
![Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human kidney shows moderate cytoplasmic positivity in cells in tubules.](http://images.novusbio.com/fullsize/antibody/nbp1-84563_rabbit-polyclonal-nt5c-antibody-immunohistochemistry-paraffin-1312202495154..png "Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human kidney shows moderate cytoplasmic positivity in cells in tubules.")
![Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human cerebral cortex shows moderate cytoplasmic positivity in neuronal cells.](http://images.novusbio.com/fullsize/antibody/nbp1-84563_rabbit-polyclonal-nt5c-antibody-immunohistochemistry-paraffin-1312202495245..png "Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human cerebral cortex shows moderate cytoplasmic positivity in neuronal cells.")
![Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human liver shows weak cytoplasmic positivity in hepatocytes as expected.](http://images.novusbio.com/fullsize/antibody/nbp1-84563_rabbit-polyclonal-nt5c-antibody-immunohistochemistry-paraffin-1312202495335..png "Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human liver shows weak cytoplasmic positivity in hepatocytes as expected.")
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![Western Blot: Goat anti-Rabbit IgG (H+L) Secondary Antibody [HRP] [NB7160] - Western blot showing vemurafenib treatment in BRAFV600E CRC cells inhibits fission mediator DRP1 with no significant effect on fusion proteins (Mfn1 & 2) using MFN-1 antibody (NBP1-51841) and corresponding secondary antibody, goat anti-rabbit IgG-HRP (NB7160). Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/33738242).](https://images.novusbio.com/images/Goat-anti-Rabbit-IgG-H+L-Secondary-Antibody-HRP-Western-Blot-NB7160-img0001.jpg "Western Blot: Goat anti-Rabbit IgG (H+L) Secondary Antibody [HRP] [NB7160] - Western blot showing vemurafenib treatment in BRAFV600E CRC cells inhibits fission mediator DRP1 with no significant effect on fusion proteins (Mfn1 & 2) using MFN-1 antibody (NBP1-51841) and corresponding secondary antibody, goat anti-rabbit IgG-HRP (NB7160). Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/33738242).")
![Flow Cytometry: Rabbit IgG Isotype Control [NBP2-24891] - Intracellular FACS analysis of mouse TLR6 polyclonal antibody (red), rabbit isotype control (green), RAW cells alone (shaded). Two micrograms of antibodies were used. Goat anti-rabbit FITC (Novus, 20302) was used as secondary.](https://images.novusbio.com/images/Rabbit--Mouse-IgG-Isotype-Control-Flow-Cytometry-NBP2-24891-img0001.jpg "Flow Cytometry: Rabbit IgG Isotype Control [NBP2-24891] - Intracellular FACS analysis of mouse TLR6 polyclonal antibody (red), rabbit isotype control (green), RAW cells alone (shaded). Two micrograms of antibodies were used. Goat anti-rabbit FITC (Novus, 20302) was used as secondary.")