重组Anti-Fatty Acid Synthase抗体[EPR7466] (ab128870)

Purified ab128870 at 1:30 dilution (2µg) immunoprecipitating Fatty Acid Synthase in HEK-293 whole cell lysate.
Lane 1 (input): HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate 10 µg
Lane 2 (+): ab128870 + HEK-293 whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab128870 in HEK-293 whole cell lysate.
VeriBlot for IP Detection Reagent (HRP)(ab131366) (1:10,000 dilution) was used for Western blotting.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: kDa

Flow Cytometry analysis of A549 (Human lung carcinoma epithelial cell) cells labelling Fatty Acid Synthase with Purified ab128870 at 1:50 dilution (10 µg/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) secondary antibody was used at 1:2000. Isotype 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 mouse liver tissue sections labeling Fatty Acid Synthase with Purified ab128870 at 1:450 dilution (1.09 µg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control. 

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat liver tissue sections labeling Fatty Acid Synthase with Purified ab128870 at 1:450 dilution (1.09 µg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control. 

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast cancer tissue sections labeling Fatty Acid Synthase with Purified ab128870 at 1:450 dilution (1.09 µg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control. 

Immunocytochemistry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Fatty Acid Synthase with Purified ab128870 at 1:50 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.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized NIH/3T3 (mouse embryonic fibroblast) cells labelling with ab128870 at 1/50 dilution, followed by ab150081 antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in NIH/3T3 cells is observed. ab195889 was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control: Secondary antibody is ab150081 at 1/1000 dilution.

Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: Fatty Acid Synthase knockout HAP1 cell lysate (20 µg)
Lane 3: A549 cell lysate (20 µg)
Lane 4: Hu liver tissue lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab128870 observed at 250 kDa. Red - loading control, ab18058, observed at 124 kDa.

ab128870 was shown to react with Fatty Acid Synthase in wild-type HAP1 cells along with additional cross reactive bands. No band was observed when Fatty Acid Synthase knockout samples were examined. Wild-type and Fatty Acid Synthase knockout samples were subjected to SDS-PAGE. ab128870 and ab18058 (loading control to Vinculin) were both diluted at 1/10,000 and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.