Anti-Apolipoprotein E antibody [EP1374Y]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

Immunohistochemical analysis of paraffin-embedded human Alzheimer's brain tissue* labelling APOE with ab52607 at 0.1ug/ml followed by a ready to use LeicaDS9800 (Bond^™ Polymer Refine Detection).

Positive staining in human Alzheimer's brain.

The section was incubated with ab52607 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond^™ Polymer Refine Detection).

Heat mediated antigen retrieval with sodium citrate buffer (pH 6.0 epitope retrieval solution1) for 20 mins.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

Intracellular 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 Fluorr^®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).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human astrocytoma tissue sections labeling Apolipoprotein E with purified ab52607 at 1/800 dilution (0.13 µg/mL). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0). The immunostaining was performed on a Leica Biosystems BOND® RX instrument. 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.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

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.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

Immunohistochemical analysis of paraffin-embedded human basal ganglia tissue* labelling APOE with ab52607 at 0.1ug/ml followed by a ready to use LeicaDS9800 (Bond^™ Polymer Refine Detection).

Positive staining in human basal ganglia.

The section was incubated with ab52607 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond^™ Polymer Refine Detection).

Heat mediated antigen retrieval with sodium citrate buffer (pH 6.0 epitope retrieval solution1) for 20 mins.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.

• IP

Lab

Immunoprecipitation - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

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

All lanes:

Immunoprecipitation - Anti-Apolipoprotein E antibody [EP1374Y] (ab52607)

Predicted band size: 36 kDa

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• IP

Lab

Immunoprecipitation - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

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

All lanes:

Immunoprecipitation - Anti-Apolipoprotein E antibody [EP1374Y] (ab52607)

Predicted band size: 36 kDa

false

• IP

Lab

Immunoprecipitation - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

Immunoprecipitation of APOE in HAP1 cells. Lysates were prepared and immunoprecipitation was performed using 2.0 µg of ab52607 pre-coupled to Protein A beads. Samples were then washed and processed for western blot

These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Immunoprecipitation - Anti-Apolipoprotein E antibody [EP1374Y] (ab52607) at 2 µg

All lanes:

HAP1 cells

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• WB

Lab

Western blot - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

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.

All lanes:

Western blot - Anti-Apolipoprotein E antibody [EP1374Y] (ab52607) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

APOE knockout HEK293T cell lysate at 20 µg

Lane 3:

HepG2 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 36 kDa

Observed band size: 36 kDa

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• WB

Lab

Western blot - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

False colour image of Western blot : Anti-Apolipoprotein E antibody [EP1374Y] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab52607 was shown to bind specifically to Apolipoprotein E. A band was observed at 34-37 kDa in wild-type HepG2 cell lysates with no signal observed at this size in APOE knockout cell line. To generate this image, wild-type and APOE knockout HepG2 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-Apolipoprotein E antibody [EP1374Y] (ab52607) at 1/1000 dilution

Lane 1:

Wild-type HepG2 cell lysate at 20 µg

Lane 2:

APOE knockout HepG2 cell lysate at 20 µg

Lane 2:

Western blot - Human APOE knockout Hep G2 cell line (<a href='/en-us/products/cell-lines/human-apoe-knockout-hep-g2-cell-line-ab280875'>ab280875</a>)

Lane 3:

Human Liver cell lysate at 20 µg

Lane 4:

Human Kidney cell lysate at 20 µg

Predicted band size: 36 kDa

Observed band size: 34 kDa,34-37 kDa

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• WB

Lab

Western blot - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

ab52607 was shown to react with APOE in wild-type HAP1 cells in Western blot with loss of signal observed in a APOE knockout cell line. Wild-type HAP1 and APOE knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab52607 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2ug/mL before imaging.

These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-Apolipoprotein E antibody [EP1374Y] (ab52607) at 1/1000 dilution

Lane 1:

Wild-type HAP1 lysate at 20 µg

Lane 2:

APOE Knockout HAP1 lysate at 20 µg

Observed band size: 36 kDa

false

• WB

Unknown

Western blot - Anti-Apolipoprotein E antibody [EP1374Y] (AB52607)

All lanes:

Western blot - Anti-Apolipoprotein E antibody [EP1374Y] (ab52607) at 1/10000 dilution

Lane 1:

Human liver lysate at 20 µg

Lane 2:

Human cerebellum lysate at 20 µg

Lane 3:

HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

Human serum lysate at 15 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 36 kDa

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