All lanes : Anti-APC antibody [EP701Y] (ab40778) at 1/5000 dilution (Purified)

Lane 1 : HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate
Lane 2 : C2C12 (Mouse myoblasts myoblast) whole cell lysate
Lane 3 : C6 (Rat glial tumor glial cell) whole cell lysate

Lysates/proteins at 15 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Observed band size: 160 kDa why is the actual band size different from the predicted?

The molecular weight observed represents the truncated APC as described in PMID: 17595655.

Blocking/Diluting buffer: 5% NFDM/TBST

Immunocytochemistry/Immunofluorescence analysis of HEK-293 (Human embryonic kidney epithelial cell) cells labeling APC with Purified ab40778 at 1:100 dilution (10 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) 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. PBS instead of the primary antibody was used as the secondary antibody only control.

Purified ab40778 at 1:70 dilution (2μg) immunoprecipitating APC in HEK-293 whole cell lysate.
Lane 1 (input): HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate 10μg
Lane 2 (+): ab40778 + HEK-293 whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab40778 in HEK-293 whole cell lysate.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: 160 kDa

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colon tissue sections labeling APC with purified ab40778 at 1/1000 dilution (1.383 µg/mL). Heat mediated antigen retrieval was performed using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

Purified ab40778 staining APC in the human cell line 293 (human embryonic kidney) by flow cytometry. Cells were fixed with 4% paraformaldehyde, permabilised with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/150. A goat anti rabbit IgG (Alexa Fluor^® 488) at a dilution of 1/2000 was used as the secondary antibody.

Isoytype control: Rabbit monoclonal IgG (Black)

Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat colon tissue sections labeling APC with purified ab40778 at 1/1000 dilution (1.383 µg/mL). Heat mediated antigen retrieval was performed using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse colon tissue sections labeling APC with purified ab40778 at 1/1000 dilution (1.383 µg/mL). Heat mediated antigen retrieval was performed using Bond™ Epitope Retrieval Solution 2 (pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

ab40778 (1:50), upurified,  staining human colon carcinoma by immunohistochemistry, paraffin-embedded tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.