Immunohistochemical analysis of human breast carcinoma tissue labeling ErbB2 / HER2 with ab237715 at 1/2000 dilution, followed by Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on the human breast carcinoma is observed. Counter stained with hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.

The section was incubated with ab237715 for 10 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

 

 

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilised SK-BR-3 (human mammary gland adenocarcinoma cell line) cells labeling ERbB 2 with ab237715 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membranous staining in SK-BR-3 cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) at 1/200 dilution (red).

Secondary antibody only control: Used PBS instead of primary antibody, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

ERbB 2 was immunoprecipitated from 0.35 mg of HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab237715 at 130 dilution. Western blot was performed from the immunoprecipitate using ab237715 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used as secondary antibody at 1/5000 dilution.

Lane 1: HeLa whole cell lysate 10 μg (Input).
Lane 2: ab237715 IP in HeLa whole lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab237715 in HeLa whole cell lysate.

Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 15 seconds.

Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methonao-permeabilized SH-BR-3 (human mammary gland adenocarcinoma cell line) cells labeling ErbB2 / HER2 with ab237715 at 1/500 dilution (red) compared with Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue).

Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077), at 1/2000 dilution was used as the secondary antibody.

Anti-ErbB2 / HER2 antibody [CAL27] (ab237715) at 1/1000 dilution + HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 137 kDa
Observed band size: 180 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

Blocking and dilution buffer: 5% NFDM/TBST.

This blot was developed using a higher sensitivity ECL substrate.

All lanes : Anti-ErbB2 / HER2 antibody [CAL27] (ab237715) at 1/1000 dilution

Lane 1 : 4T1 (mouse mammary gland carcinoma cell line) whole cell lysate
Lane 2 : C6 (rat glial tumor glial cell) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/5000 dilution

Predicted band size: 137 kDa
Observed band size: 180 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

Blocking and dilution buffer: 5% NFDM/TBST.

This blot was developed using a higher sensitivity ECL substrate.

Formalin-fixed, paraffin-embedded human breast carcinoma tissue stained for ErbB2 / HER2 using ab237715  at 0.3 μg/ml in immunohistochemical analysis.