NT5C Antibody

Simple Western: NT5C Antibody [NBP1-84563]

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).

Knockdown Validated: NT5C Antibody [NBP1-84563]

Knockdown Validated: NT5C Antibody [NBP1-84563] - NT5C regulates cell motility and cell spreading. NT5C regulates single cell spreading. HT1080 cells stably expressing siRNA were trypsinized, seeded onto fibronectin coated plates and fixed at 45 min post-seeding. (left) Cell area was determined manually in ImageJ and is expressed relative to siControl cells. The experiment was repeated 3 times independently. n = 326-431 cells per cell line; Error bars are sem; *p < 0.05, **p < 0.01. (top right) Representative image. (bottom right) Representative immunoblot from experimental cells. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/srep39985), licensed under a CC-BY license.

Western Blot: NT5C Antibody [NBP1-84563] -

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 (https://pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: NT5C Antibody [NBP1-84563] -

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 (https://pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: NT5C Antibody [NBP1-84563] -

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 (https://pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: NT5C Antibody [NBP1-84563] -

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 (https://pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: NT5C Antibody [NBP1-84563] -

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 (https://pubmed.ncbi.nlm.nih.gov/28059163), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -

Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human intestine shows strong cytoplasmic positivity in glandular cells.

Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -

Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human kidney shows moderate cytoplasmic positivity in cells in tubules.

Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -

Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human cerebral cortex shows moderate cytoplasmic positivity in neuronal cells.

Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -

Immunohistochemistry-Paraffin: NT5C Antibody [NBP1-84563] -Staining of human liver shows weak cytoplasmic positivity in hepatocytes as expected.