Anti-Src (phospho Y529) 抗体 [Y232] (ab32078)

WB analysis. Lane 1:Untreated HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysates 15ug and Lane 2:HeLa (Human epithelial cell line from cervix adenocarcinoma) treated with H₂O₂ at 10mM for 1 h. whole cell lysates 15ug and Lane 3:HeLa (Human epithelial cell line from cervix adenocarcinoma) treated with H₂O₂ at 10mM for 1 h. whole cell lysates 15ug. Then the membrane was incubated with phosphatase. Primary ab used at 1:30,000 dilution. 5% NFDM/TBST  was used as Blocking buffer and Diluting buffer. Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) was used as a Secondary ab at 1:20,000 dilution. The Exposure time was 30 seconds.

Immunocytochemistry/Immunofluorescence analysis of HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labelling Src (phospho Y529) with ab32078 at 1/500. Cells were fixed with 4% Paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. Cells were counter-stained with ab195889 Anti-Alpha Tubulin antibody [DM1A] (1/200) - Microtubule Marker (Alexa Fluor^® 594). DAPI (blue) was used as a nuclear counterstain.

The green staining was increased in the H2O2 (10mM, 1 hour) treated HeLa cells when compared with HeLa cells without treatment. After LP treatment, the green signaling was obviously decreased.

For the pan antibody, there was no great difference after H₂O₂ (10 mM, 1 hour) or EGF (100 ng/mL, 10 minutes) + LP treatment.

The data showed mostly Cytoplasm and Membran staining for Ab32078.

Dot Blot analysis of Lane 1: Src (pY529) phospho peptide and Lane 2: Src non-phospho peptide labeling Src (phospho Y529) with ab32078 at 1/1000 dilution. 5% NFDM/TBST was used as the diluting and blocking buffer. ab97051 Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated was used as the secondary antibody at 1/100000 dilution. Exposure time: 3 minutes.