Anti-Integrin alpha 5 抗体 (ab112183)

Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: Integrin alpha 5 knockout HAP1 whole cell lysate (20 µg)
Lane 3: Jeg3 whole cell lysate (20 µg)
Lane 4: HeLa whole cell lysate (20 µg)

Lanes 1 - 4: Merged signal (red and green). Green - ab112183 observed at 140 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab112183 was shown to recognize Integrin alpha 5 when Integrin alpha 5 knockout samples were used, along with additional cross-reactive bands. Wild-type and Integrin alpha 5 knockout samples were subjected to SDS-PAGE. Ab112183 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1000 dilution and 1/10000 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 1/10000 dilution for 1 hour at room temperature before imaging.

 

IHC image of Integrin alpha 5 staining in Human normal placenta formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab112183, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

All lanes : Anti-Integrin alpha 5 antibody (ab112183) at 1 µg/ml

Lane 1 : Human placenta tissue lysate - total protein (ab29745)
Lane 2 : JEG-3 (Human placental choriocarcinoma cell line) Whole Cell Lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

Developed using the ECL technique.

Performed under reducing conditions.

Predicted band size: 114 kDa
Observed band size: 145 kDa (why is the actual band size different from the predicted?)
Additional bands at: 32 kDa, 50 kDa, 60 kDa. We are unsure as to the identity of these extra bands.


Exposure time: 30 seconds


Integrin alpha 5 contains an extensive number of potential glycosylation sites (SwissProt) which may explain its migration at a higher molecular weight than predicted.