All lanes : Anti-GALT antibody [EPR12555] (ab178406) at 1/1000 dilution

Lane 1 : Wild-type HeLa cell lysate
Lane 2 : GALT knockout HeLa cell lysate
Lane 3 : HepG2 cell lysate
Lane 4 : A549 cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 43 kDa

Lanes 1-4: Merged signal (red and green). Green - ab178406 observed at 43 kDa. Red - loading control ab7291 observed at 50 kDa.

 ab178406 Anti-GALT antibody [EPR12555] was shown to specifically react with Galactose-1-Phosphate Uridylyltransferase in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265027 (knockout cell lysate ab257958) was used. Wild-type and Galactose-1-Phosphate Uridylyltransferase knockout samples were subjected to SDS-PAGE. ab178406 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.

 

Lanes 1-2 : Anti-GALT antibody [EPR12555] (ab178406) at 1/1000 dilution
Lanes 3-4 : Anti-GAPDH antibody [6C5] - Loading Control (ab8245) at 1/2000 dilution

Lanes 1 & 3 : GALT knockout HAP1 cell lysate
Lanes 2 & 4 : Wild-type HAP1 cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 43 kDa

 

Lanes 1 and 2: Green signal from target – ab178406 observed at 43 kDa

Lanes 3 and 4: Red signal from loading control – ab8245 observed at 37 kDa

Lanes 5 and 6: Merged (red and green) signal

ab178406 was shown to react with GALT when GALT knockout samples were used, along with additional cross-reactive bands. Wild-type and GALT knockout samples were subjected to SDS-PAGE. ab178406 and ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/2000 respectively and incubated overnight at 4ºC. Blots were developed withGoat anti-Rabbit IgG H&L (IRDye® 800CW)preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776)secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.

All lanes : Anti-GALT antibody [EPR12555] (ab178406) at 1/1000 dilution

Lane 1 : K562 cell lysate
Lane 2 : MDA-MB-435 cell lysate
Lane 3 : Human fetal brain lysate
Lane 4 : Human fetal liver lysate
Lane 5 : HepG2 cell lysate

Lysates/proteins at 10 µg per lane.

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

Developed using the ECL technique.

Predicted band size: 43 kDa

Western blot analysis on Immunoprecipitation pellet from either 1) Human fetal liver lysate, or 2) 1xPBS (negative control); showing GALT, immunoprecipitated using ab178406 at 1/10 dilution with HRP-conjugated anti-rabbit IgG preferentially detecting the non-reduced form of rabbit IgG.