All lanes : Anti-CD147 antibody [EPR4053] (ab108308) at 1/1000 dilution

Lane 1 : Wild-type A549 cell lysate
Lane 2 : BSG knockout A549 cell lysate
Lane 3 : Raji cell lysate

Lysates/proteins at 30 µg per lane.

Performed under reducing conditions.

Predicted band size: 42 kDa
Observed band size: 42-70 kDa why is the actual band size different from the predicted?

Lanes 1 - 3: Merged signal (red and green). Green - ab108308 observed at 42-70 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A] observed at 55kDa.

ab108308 was shown to react with CD147 in wild-type A549 cells in western blot with loss of signal observed in BSG knockout cell line ab273748 (knockout cell lysate ab275500). Wild-type and BSG knockout A549 cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab108308 and ab7291 (Mouse anti-Alpha Tubulin [DM1A] overnight at 4°C at a 1 in 1000 Dilution and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 hour at room temperature before imaging.

Immunocytochemistry/ Immunofluorescence analysis of HeLa (human cervix adenocarcinoma epithelial cell) cells labeling CD147 with purified ab108308 at 1/500 dilution (5 µg/mL). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at 1/200 (2.5 µg/mL). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/mL) dilution. DAPI (blue) was used as nuclear counterstain. ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 (2.5 µg/mL) was used as the secondary antibody only control.

ab108308, at 1/250, staining CD147 in human glioma tissue by immunohistochemistry.

Immunohistochemical analysis of paraffin-embedded human glioma tissue labelling CD147 with ab108308 at 1/250 dilution. Heat mediated antigen retrieval with citrate buffer, pH 6 was performed.

All lanes : Anti-CD147 antibody [EPR4053] (ab108308) at 1/1000 dilution

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : BSG knockout HEK293T cell lysate
Lane 3 : HeLa cell lysate
Lane 4 : U-87 MG 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

Performed under reducing conditions.

Predicted band size: 42 kDa
Observed band size: 50 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab108308 observed at 50 kDa. Red - loading control ab8245 observed at 36 kDa.

ab108308 Anti-CD147 antibody [EPR4053] was shown to specifically react with CD147 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266331 (knockout cell lysate ab256853) was used. Wild-type and CD147 knockout samples were subjected to SDS-PAGE. ab108308 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 : Anti-CD147 antibody [EPR4053] (ab108308) at 1/1000 dilution

Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : BSG (Basigin) knockout HAP1 whole cell lysate
Lane 3 : HeLa whole cell lysate
Lane 4 : U-87 MG whole cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 42 kDa

Lanes 1 - 4: Merged signal (red and green). Green - ab108308 observed at 42 kDa. Red - loading control, ab18058, observed at 130 kDa.

ab108308 was shown to specifically react with Basigin in wild-type HAP1 cells as signal was lost in BSG (Basigin) knockout cells. Wild-type and BSG (Basigin) knockout samples were subjected to SDS-PAGE. Ab108308 and ab18058 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/1000 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-CD147 antibody [EPR4053] (ab108308) at 1/1000 dilution

Lane 1 : Jurkat cell lysate
Lane 2 : HuT-78 cell lysate
Lane 3 : A431 cell lysate

Lysates/proteins at 10 µg per lane.

Performed under reducing conditions.

Predicted band size: 42 kDa

Anti-CD147 antibody [EPR4053] (ab108308) at 1/1000 dilution + HeLa cell lysate at 10 µg

Performed under reducing conditions.

Predicted band size: 42 kDa

Immunohistochemical analysis of paraffin-embedded human brain tissue labelling CD147 with ab108308 at 1/250 dilution. Heat mediated antigen retrieval with citrate buffer, pH 6 was performed.