Immunocytochemistry - Drosha antibody (ab12286)

Human female amniocytes immunostained with ab12286 Drosha (FITC) (1/25 dilution). The DNA is labelled red with propidium iodide. This image was submitted as part of a review by Ahmad Khalil.

Ahmad M. Khalil and Daniel J. Driscoll, University of Florida College of Medicine Genetics Institute.

Immunocytochemistry/ Immunofluorescence - Drosha antibody (ab12286)

ab12286 at a 1/25 dilution staining human HeLa cells by immunocytochemistry. The antibody was incubated with the cells for 1 hour and then detected using a Cy3 conjugated donkey anti-rabbit polyclonal antibody.

This image is courtesy of an Abreview submitted on 7 February 2006.

Western blot - Drosha antibody (ab12286)

All lanes : Anti-Drosha antibody - ChIP Grade (ab12286) at 1 µg/ml

Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
Lane 2 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate

Lysates/proteins at 20 µg per lane.

Secondary
Goat polyclonal to Rabbit IgG (Alexa Fluor® 680) at 1/15000 dilution

Performed under reducing conditions.

Predicted band size : 159 kDa
Observed band size : 170 kDa (why is the actual band size different from the predicted?)

Western blot - Anti-Drosha antibody - ChIP Grade (ab12286)

All lanes : Anti-Drosha antibody - ChIP Grade (ab12286) at 1 µg/ml

Lane 1 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate
Lane 2 : NIH 3T3 (Mouse embryonic fibroblast cell line) Nuclear Lysate

Lysates/proteins at 20 µg per lane.

Secondary
Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (HRP), pre-adsorbed (ab97080) at 1/5000 dilution
developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 159 kDa
Observed band size : 170 kDa (why is the actual band size different from the predicted?)
Additional bands at : 115 kDa. We are unsure as to the identity of these extra bands.

Exposure time : 12 minutes

Abcam recommends using milk as the blocking agent. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above.

Immunocytochemistry/ Immunofluorescence - Drosha antibody - ChIP Grade (ab12286)

ICC/IF image of ab12286 stained HepG2 cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab12286, 1µg/ml) overnight at +4ºC. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

Immunocytochemistry/ Immunofluorescence - Drosha antibody - ChIP Grade (ab12286)

ICC/IF image of ab12286 stained Hela cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab12886, 5µg/ml) overnight at +4ºC. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.