All lanes : 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
Lane 2 : NDRG1 knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lane 3 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 4 : Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

Predicted band size: 43 kDa
Observed band size: 43 kDa

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.

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.

All lanes : Anti-NDRG1 antibody [EPR5593] (ab124689) at 1/10000 dilution

Lane 1 : Wild-type HEK-293 whole cell lysate
Lane 2 : NDRG1 knockout HEK-293 whole cell lysate
Lane 3 : Jurkat whole cell lysate
Lane 4 : LnCap whole cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 43 kDa

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.

All lanes : Anti-NDRG1 antibody [EPR5593] (ab124689) at 0.0083 µg/ml

Lane 1 : Mouse brain lysate
Lane 2 : Rat brain lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 43 kDa


Exposure time: 3 minutes

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 .

Equilibrium disassociation constant (K_D)
Learn more about K_D

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All lanes : Anti-NDRG1 antibody [EPR5593] (ab124689) at 1/10000 dilution

Lane 1 : CaCo-2 (Human colorectal adenocarcinoma cell line) lysate
Lane 2 : Human fetal kidney lysate
Lane 3 : LnCaP (Human prostate cancer cell line) lysate
Lane 4 : HeLa (Human epithelial cell line from cervix adenocarcinoma) lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat-anti-rabbit HRP at 1/2000 dilution

Predicted band size: 43 kDa

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

ab124689 showing positive staining in human cervical carcinoma tissue.

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

ab124689 showing positive staining in human colonic adenocarcinoma tissue.

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

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.

ab124689 showing positive staining in human breast carcinoma tissue.

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