All lanes : Anti-MVP antibody [EPR23594-106] (ab273093) at 1/1000 dilution

Lane 1 : Wild-type HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate with Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS
Lane 2 : MVP knockout HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate with Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS
Lane 3 : Caco-2 (human colorectal adenocarcinoma epithelial cell) whole cell lysate with Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS

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: 99 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?

Lanes 1-3: Merged signal (red and green). Green - ab273093 observed at 110 kDa. Red - loading control ab8245 (Mouse monoclonal [6C5] to GAPDH) observed at 36 kDa.

ab273093 Anti-MVP antibody [EPR23594-106] was shown to specifically react with MVP in wild-type HeLa cells in Western blot. Significant decrease (8.7 % of intensity compared to the WT band) of signal was observed when MVP knockout cell line ab264817 (knockout cell lysate ab257544) was used.

ab273093 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at 4? overnight 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 10000 dilution for 1 hour at room temperature before imaging.

All lanes : Anti-MVP antibody [EPR13226(B)] (ab177145) at 1/1000 dilution

Lane 1 : Wild-type HeLa lysate
Lane 2 : MVP knockout HeLa lysate
Lane 3 : A549 lysate
Lane 4 : MOLT-4 lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 99 kDa

Lanes 1-4: Merged signal (red and green). Green - ab177145 observed at 99 kDa. Red - loading control ab8245 observed at 37 kDa.

 ab177145 Anti-MVP antibody [EPR13226(B)] was shown to specifically react with MVP in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264817 (knockout cell lysate ab257544) was used. Wild-type and MVP knockout samples were subjected to SDS-PAGE. ab177145 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 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-MVP antibody [EPR13227(B)] (ab175239) at 1/2000 dilution

Lane 1 : Wild-type HeLa lysate
Lane 2 : MVP knockout HeLa lysate
Lane 3 : A549 lysate
Lane 4 : MOLT-4 lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 99 kDa

Lanes 1-4: Merged signal (red and green). Green - ab175239 observed at 110 kDa. Red - loading control ab8245 observed at 37 kDa.

 ab175239 Anti-MVP antibody [EPR13227(B)] was shown to specifically react with MVP in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264817 (knockout cell lysate ab257544) was used. Wild-type and MVP knockout samples were subjected to SDS-PAGE. ab175239 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 2000 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: 17 bp deletion in exon 6.

 

Allele-2: 1 bp deletion in exon 6.