All lanes : Anti-MDH2 antibody [EPR14883(B)] (ab181857) at 1/1000 dilution

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : MDH2 knockout HEK293T cell lysate
Lane 3 : K-562 cell lysate
Lane 4 : Human eyeball tissue 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: 36 kDa
Observed band size: 36 kDa

Lanes 1-4: Merged signal (red and green). Green - ab181857 observed at 36 kDa. Red - loading control ab7291 observed at 50 kDa.

 ab181857 Anti-MDH2 antibody [EPR14883(B)]  was shown to specifically react with MDH2 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266449 (knockout cell lysate ab257533) was used. Wild-type and MDH2 knockout samples were subjected to SDS-PAGE. ab181857 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) 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 : Anti-MDH2 antibody [EPR14882(B)] (ab181873) at 1/1000 dilution

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : MDH2 knockout HEK293T cell lysate
Lane 3 : K-562 cell lysate
Lane 4 : Human eyeball tissue 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: 36 kDa
Observed band size: 36 kDa

Lanes 1-4: Merged signal (red and green). Green - ab181873 observed at 36 kDa. Red - loading control ab7291 observed at 50 kDa.

 ab181873 Anti-MDH2 antibody [EPR14882(B)]  was shown to specifically react with MDH2 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266449 (knockout cell lysate ab257533) was used. Wild-type and MDH2 knockout samples were subjected to SDS-PAGE. ab181873 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) 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.

Allele-1: 4 bp deletion in exon 4

 

Allele-2: Insertion of the selection cassette in exon 4.

 

Representative images of MDH2 knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using an EVOS M5000 microscope.