Western blot - Anti-HIF1 alpha antibody [H1alpha67] - ChIP Grade (ab1)

All lanes : Anti-HIF1 alpha antibody [H1alpha67] - ChIP Grade (ab1) at 5 µg/ml

Lane 1 : Hela - Vehicle only (Negative Control) Whole Cell Lysate
Lane 2 : Hela - DFO treated (0.5mM, 24h) Whole Cell Lysate

Lysates/proteins at 25 µg per lane.

Secondary
Goat polyclonal Secondary Antibody to Mouse IgG - H&L (HRP), pre-adsorbed (ab97040) at 1/5000 dilution
developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 92 kDa
Observed band size : 120 kDa (why is the actual band size different from the predicted?)
Additional bands at : 60 kDa. We are unsure as to the identity of these extra bands.

Exposure time : 20 minutes

Western blot - Anti-HIF1 alpha antibody [H1alpha67] - ChIP Grade (ab1)

Anti-HIF1 alpha antibody [H1alpha67] - ChIP Grade (ab1) at 1/400 dilution + Human Cell lysate - whole cell (human lung adenocarcinoma cell line ADLC-5M2) treated for 16 hours with 100 micromolar deferoxamine (DFO) at 20 µg

Performed under reducing conditions.

Predicted band size : 92 kDa
Observed band size : 120 kDa (why is the actual band size different from the predicted?)


PVDF membrane was used and blocked for 16 hours in 5% milk.

This image is taken from an Abreview submitted by Mike Campa, no further information is known about this image

Immunocytochemistry/ Immunofluorescence - HIF1 alpha antibody [H1alpha67] (ab1)

ab1 at 1/200 dilution staining HIF1 alpha in human 293FT cells by Immunocytochemistry/ Immunofluorescence. Cells were fixed in formaldehyde, permeabilized in 0.5% Trition X-100 and blocked in 5% BSA for 1 hour at 25°C. The primary antibody was used at 1/200 dilution in PBS and incubated with sample at 4°C for 12 hours. An Alexa Fluor® 488 conjugated Goat polyclonal to mouse IgG was used as secondary at 1/500 dilution.

This Image is courtesy of an anonymous Abreview

Immunohistochemistry (Frozen sections) - HIF1 alpha antibody [H1alpha67] (ab1)

ab1 staining HIF1 alpha in mouse liver tissue section by Immunohistochemistry (Frozen sections). Tissue samples were fixed with formaldehyde and permeablized with 0.2% Triton-X100 before blocking with 2% BSA for 30 minutes at 20°C. The sample was incubated with primary antibody (1/200) for 9 hours at 4°C. An Alexa Fluor®555-conjugated Goat polyclonal to mouse IgG was used as secondary antibody at 1/200 dilution. DAPI was used to stain the cell nuclei (blue).

This image is courtesy of an anonymous Abreview