Anti-NDRG1 antibody [EPR5593]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NDRG1 antibody [EPR5593] (AB124689)

Immunohistochemical analysis of Paraffin-embedded sections human liver carcinoma tissue labelling NDRG1 with ab124689 at 1/1000 dilution, followed by a ready to use secondary Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Staining on human liver carcinoma tissue is observed. Counter stained with Haematoxylin. Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NDRG1 antibody [EPR5593] (AB124689)

ab124689 showing positive staining in human normal kidney tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NDRG1 antibody [EPR5593] (AB124689)

Immunohistochemical analysis of Paraffin-embedded sections human colon tissue labelling NDRG1 with ab124689 at 1/1000 dilution, followed by a ready to use secondary Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Staining on human colon tissue is observed. Counter stained with Haematoxylin. Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-NDRG1 antibody [EPR5593] (AB124689)

Intracellular Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling NDRG1 with purified ab124689 at 1/20 dilution (5 ug/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150081) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as a isotype control. Cell without incubation with primary antibody and secondary antibody (Blue) were used as unlabeled control.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NDRG1 antibody [EPR5593] (AB124689)

ab124689 at 1/100 dilution, staining NDRG1 in Formalin fixed Paraffin-embedded human colon tissue by Immunohistochemistry.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-NDRG1 antibody [EPR5593] (AB124689)

Immunocytochemistry/ Immunofluorescence analysis of Jurkat (Human T cell leukemia T lymphocyte) cells labeling NDRG1 using ab124689. The cells were fixed with 100% Methanol then permeabilized with 0.1% Triton X-100. The cells were then incubated with ab124689 at 1 : 50 dilution followed by a further incubation with a Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI. Cells were counterstained using ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1 : 200 dilution (shown in red). Secondary antibody only control : PBS instead of the primary antibody.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-NDRG1 antibody [EPR5593] (AB124689)

ab124689, at 1/100 dilution, staining NDRG1 in HeLa (Human epithelial cell line from cervix adenocarcinoma) cells by Immunofluorescence.

• IP

Lab

Immunoprecipitation - Anti-NDRG1 antibody [EPR5593] (AB124689)

Lane 1 (input) : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate 10μg.

Lane 2 (+) : ab124689 & HeLa whole cell lysate
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab124689 in HeLa whole cell lysate

ab124689 (Purified) at 1/50 dilution (20μg/ml) immunoprecipitating NDRG1 in HeLa whole cell lysate. For western blotting, ab124689 at 1/500 dilution (1.86 μg/mL) and VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1/1000 dilution.

Blocking and diluting buffer : 5% NFDM /TBST .

All lanes:

Immunoprecipitation - Anti-NDRG1 antibody [EPR5593] (ab124689)

Predicted band size: 43 kDa

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• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NDRG1 antibody [EPR5593] (AB124689)

Immunohistochemical analysis of Paraffin-embedded sections rat colon tissue labelling NDRG1 with ab124689 at 1/1000 dilution, followed by a ready to use secondary Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Staining on rat colon tissue is observed. Counter stained with Haematoxylin. Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NDRG1 antibody [EPR5593] (AB124689)

Immunohistochemical analysis of Paraffin-embedded sections mouse colon tissue labelling NDRG1 with ab124689 at 1/1000 dilution, followed by a ready to use secondary Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Staining on mouse colon tissue is observed. Counter stained with Haematoxylin. Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• WB

Lab

Western blot - Anti-NDRG1 antibody [EPR5593] (AB124689)

Lanes 1 - 4 : Merged signal (red and green). Green - ab124689 observed at 43 kDa. Red - loading control, ab130007, observed at 130 kDa.

ab124689 was shown to recognize in wild-type HEK-293 cells as signal was lost at the expected MW in NDRG1 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and NDRG1 knockout samples were subjected to SDS-PAGE. ab124689 and ab130007 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/10000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

Lanes 1 - 4:

Western blot - Anti-NDRG1 antibody [EPR5593] (ab124689) at 1/10000 dilution

Lane 1:

Western blot - Alexa Fluor® 647 Anti-NDRG1 antibody [EPR5593] (<a href='/en-us/products/primary-antibodies/alexa-fluor-647-ndrg1-antibody-epr5593-ab199471'>ab199471</a>)

Lane 1:

Wild-type HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

Lane 2:

NDRG1 knockout HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

Lane 2:

Western blot - Human NDRG1 knockout HEK-293 cell line (<a href='/en-us/products/cell-lines/human-ndrg1-knockout-hek-293-cell-line-ab261856'>ab261856</a>)

Lane 3:

Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate at 20 µg

Lane 4:

LNCaP (Human prostate cancer cell line) whole cell lysate at 20 µg

Predicted band size: 43 kDa

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• WB

Lab

Western blot - Anti-NDRG1 antibody [EPR5593] (AB124689)

Lanes 1-4 : Merged signal (red and green). Green - ab124689 observed at 43 kDa. Red - loading control ab8245 observed at 36 kDa.

ab124689 Anti-NDRG1 antibody [EPR5593] was shown to specifically react with NDRG1 in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab267301 (knockout cell lysate ab257551) was used. Wild-type and NDRG1 knockout samples were subjected to SDS-PAGE. ab124689 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-NDRG1 antibody [EPR5593] (ab124689) at 1/1000 dilution

Lane 1:

Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg

Lane 2:

NDRG1 knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg

Lane 2:

Western blot - Human NDRG1 knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-ndrg1-knockout-hek-293t-cell-line-ab267301'>ab267301</a>)

Lane 3:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 4:

Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 43 kDa

Observed band size: 43 kDa

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• WB

Supplier Data

Western blot - Anti-NDRG1 antibody [EPR5593] (AB124689)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. LY294002 treatment decreased the expression of phospho T346 NDRG1(PMID : 18787837). In Western blot, anti-NDRG1 antibody (ab124689) Pan control staining at 1/1000 dilution. In Western blot, anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution. Alkaline phosphatase was used to treated membrane. Exposure time : 103 seconds.

All lanes:

Western blot - Anti-NDRG1 (phospho T346) antibody [EPR26325-247] (<a href='/en-us/products/primary-antibodies/ndrg1-phospho-t346-antibody-epr26325-247-ab307706'>ab307706</a>) at 1/1000 dilution

Lane 1:

HeLa starved for 4 hours, whole cell lysate at 20 µg

Lane 2:

HeLa starved for 4 hours, then treated with 50uM LY294002 for 4 hours, whole cell lysate at 20 µg

Lane 3:

HeLa starved for 4 hours, whole cell lysate (phosphatase treated membrane)

Lane 4:

HeLa starved for 4 hours, then treated with 50uM LY294002 for 4 hours, whole cell lysate (phosphatase treated membrane)

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/5000 dilution

Observed band size: 48 kDa

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Exposure time: 103s

• WB

Lab

Western blot - Anti-NDRG1 antibody [EPR5593] (AB124689)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. ab181602 was used as GAPDH loading control.

All lanes:

Western blot - Anti-NDRG1 antibody [EPR5593] (ab124689) at 1/10000 dilution

Lane 1:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

Mouse brain lysate at 20 µg

Lane 3:

Rat brain lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 43 kDa

Observed band size: 48 kDa

false

• WB

Supplier Data

Western blot - Anti-NDRG1 antibody [EPR5593] (AB124689)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. Insulin treatment increased the expression of phospho T346 NDRG1 (PMID : 18787837). In Western blot, anti-NDRG1 antibody (ab124689) Pan control staining at 1/1000 dilution. In Western blot, anti- Vinculin antibody (ab129002) loading control staining at 1/10000 dilution. Exposure time : Lane 1 and 2 : 70 seconds, Lane 3 and 4 : 48 seconds.

All lanes:

Western blot - Anti-NDRG1 (phospho T346) antibody [EPR26325-247] (<a href='/en-us/products/primary-antibodies/ndrg1-phospho-t346-antibody-epr26325-247-ab307706'>ab307706</a>) at 1/1000 dilution

Lane 1:

C2C12 whole cell lysate at 20 µg

Lane 2:

C2C12 treated with 20nM insulin, whole cell lysate at 20 µg

Lane 3:

Rat-1 starved for 4 hours, whole cell lysate at 20 µg

Lane 4:

Rat-1 starved for 4 hours, then treated with 20nM insulin, whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/5000 dilution

Observed band size: 48 kDa

false

Exposure time: 70s

• WB

CiteAb

Western blot - Anti-NDRG1 antibody [EPR5593] (AB124689)

NDRG1 western blot using anti-NDRG1 antibody [EPR5593] ab124689. Publication image and figure legend from Tang, M. K., Liang, Y. J., et al., 2013, PLoS One, PubMed 23555679.

ab124689 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab124689 please see the product overview.

Silencing NDRG1 in PML+/+ MEFs alters gene expression.Semi-quantitative RT-PCR analysis revealed that silencing NDRG1 inhibited Ndrg1, Pml and p53 expression. β-actin served as an internal control for normalization.

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