Immunocytochemistry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling Apolipoprotein E with Purified ab52607 at 1:50 dilution (2.0 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.

All lanes : Anti-Apolipoprotein E antibody [EP1374Y] (ab52607) at 1/1000 dilution (Unpurified)

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : APOE knockout HEK293T cell lysate
Lane 3 : HepG2 cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

Predicted band size: 36 kDa
Observed band size: 36 kDa

Lanes 1-3: Merged signal (red and green). Green - ab52607 observed at 36 kDa. Red - loading control ab7291 observed at 50 kDa. 

 ab52607 Anti-Apolipoprotein E antibody [EP1374Y] was shown to specifically react with Apolipoprotein E in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266546 (knockout cell lysate ab256838) was used.  Wild-type and Apolipoprotein E knockout samples were subjected to SDS-PAGE.  ab52607 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated at room temperature for 2. 5 hours at 1 in 1000 dilution and 1 in 20000 dilution respectively.  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 in 20000 dilution for 1 hour at room temperature before imaging.

Flow cytometry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling Apolipoprotein E (red) with purified ab52607 (Unpurified) at a 1/250 dilution (10ug/mL). Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A goat anti rabbit IgG (Alexa Fluor^®488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (Black) (ab172730). Blue (unlabeled control) - Cell without incubation with primary antibody and secondary antibody (Blue).

Purified ab52607 at 1/20 dilution (0.5µg) immunoprecipitating Apolipoprotein E in HepG2 whole cell lysate.
Lane 1 (input): HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate 10µg
Lane 2 (+): ab52607 + HepG2 whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab52607 in HepG2 whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/5000 dilution) was used for Western blotting.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: 36 kDa

Purified ab52607 at 1/20 dilution (0.5µg) immunoprecipitating Apolipoprotein E in Human serum.
Lane 1 (input): Human serum 10µg
Lane 2 (+): ab52607 + Human serum.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab52607 in Human serum.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: 36 kDa