Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - N Cadherin antibody (ab12221)

Paraffin-embedded sections of 15-day mouse embryo. Antigen retrieval: microwave treatment, 3 minutes x 10 times, 10 mM citrate buffer. Primary antibody: 10 ug/ml ab12221, 1 hour, room temperature. Secondary antibody: Envision plus Colorimetric detection: DAB

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - N Cadherin antibody (ab12221)

 ab12221 at 1/200 dilution staining rat heart tissue sections by IHC-P. The tissue was fixed with Histochoice and paraffin sectioned prior to staining. An Alexa-Fluor conjugated donkey anti-rabbit antibody was used as the secondary.

The image depicts stained N-Cadherin (green) and counterstained nuclei (DAPI-blue) overlay (left panel). The DIC (phase) image of the same tissue, superimposed with fluorescence overlay (right panel) confirms the localization of the N-Cadherin at intercalated discs region. Note, the phase contrast image shows typical uninucleated, cylindrical morphology of cardiac myocytes; that are often branched and joined to each other end-to-end (intercalated discs) to form interlacing network.

This image is courtesy of an Abreview submitted by Dr Mal Niladri

Immunocytochemistry/ Immunofluorescence - N Cadherin antibody (ab12221)

ab12221, at 1/100, staining N Cadherin in mouse differentiated embryonic stem cells by immunocytochemistry / immunofluorescence. Cells were methanol/acetone (1/1) fixed and permeabilized in 0.1% triton X prior to blocking in 1% serum and 0.1% BSA for 30 minutes at 21ºC. Alexa fluor® 488 goat polyclonal to rabbit Ig, diluted 1/200, was used as the secondary antibody.

This image is courtesy of an Abreview from Sarah Ritson.

Western blot - N Cadherin antibody (ab12221)

All lanes : Anti-N Cadherin antibody (ab12221) at 1/100 dilution

Lane 1 : MDA-MB-231 cells (Negative Control)
Lane 2 : U2OS cells (Positive Control)

Lysates/proteins at 200000 cells per lane.

Secondary
Goat anti-rabbit HRP at 1/2000 dilution
developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 99.9 kDa
Observed band size : 100 kDa (why is the actual band size different from the predicted?)
Additional bands at : 75 kDa. We are unsure as to the identity of these extra bands.

Exposure time : 5 minutes

This image is courtesy of an anonymous Abreview

Western blot - N Cadherin antibody (ab12221)

All lanes : Anti-N Cadherin antibody (ab12221) at 1/100 dilution

Lane 1 : Whole cell lysate prepared from human glioblastoma cell line U373MG
Lane 2 : Whole cell lysate prepared from human glioblastoma cell line U251

Lysates/proteins at 100 µg per lane.

Secondary
HRP monoclonal anti-rabbit immunoglobulins at 1/7500 dilution
developed using the ECL technique

Predicted band size : 99.9 kDa
Observed band size : 100 kDa (why is the actual band size different from the predicted?)


Exposure time : 10 minutes

Distinct bands were found in all tested cell lines with no unspecific bands.

Image courtesy of an anonymous Abreview.