Anti-ENT1 antibody [SP120]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ENT1 antibody [SP120] (AB182023)

Immunohistochemical analysis of formalin-fixed paraffin-embedded Human pancreas tissue labeling ENT1 with ab182023.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ENT1 antibody [SP120] (AB182023)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human pancreas tissue sections labeling ENT1 with Purified ab182023 at 1/100 dilution (0.77 µg/ml). Heat mediated antigen retrieval was performed Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins. 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

Collaborator

Immunocytochemistry - Anti-ENT1 antibody [SP120] (AB182023)

ab182023 was shown to react with SLC29A1 in wild-type HAP1 cells in Immunocytochemistry with loss of signal observed in a SLC29A1 knockout cell line. Wild-type and knockout cells were mixed and pelleted at a 1 : 1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with . The cells were then incubated with ab182023 at 1/200 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat anti-rabbit secondary antibody to (Alexa Fluor^® 555) at 0.5 μg/ml. Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880). 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.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ENT1 antibody [SP120] (AB182023)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human colon carcinoma tissue sections labeling ENT1 with Purified ab182023 at 1/100 dilution (0.77 µg/ml). Heat mediated antigen retrieval was performed Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins. 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.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-ENT1 antibody [SP120] (AB182023)

Flow Cytometry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling ENT1 with purified ab182023 at (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody. Isotype control - Rabbit monoclonal IgG (ab172730) / Black. Unlabeled control - Unlabelled cells / blue.

• WB

Collaborator

Western blot - Anti-ENT1 antibody [SP120] (AB182023)

ab182023 was shown to react with SLC29A1 in wild-type HAP1 cells in Western blot with loss of signal observed in a SLC29A1 knockout cell line. Wild-type HAP1 and SLC29A1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab182023 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with goat anti-rabbit HRP 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-ENT1 antibody [SP120] (ab182023) at 1/1000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 20 µg

Lane 2:

SLC29A1 knockout HAP1 cell lysate at 20 µg

Predicted band size: 50 kDa

false

• WB

Lab

Western blot - Anti-ENT1 antibody [SP120] (AB182023)

False colour image of Western blot : Anti-ENT1 antibody [SP120] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab182023 was shown to bind specifically to ENT1. A band was observed at 40-60 kDa in wild-type HEK-293T cell lysates with no signal observed at this size in SLC29A1 knockout cell line. To generate this image, wild-type and SLC29A1 knockout HEK-293T cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-ENT1 antibody [SP120] (ab182023) at 1/1000 dilution

Lane 1:

Wild-type HEK-293T cell lysate at 20 µg

Lane 2:

SLC29A1 knockout HEK-293T cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

HAP1 cell lysate at 20 µg

Lane 5:

HEK-293 cell lysate at 20 µg

Lane 6:

U-87 MG cell lysate at 20 µg

Secondary

Lanes 1 - 6:

Goat anti-Rabbit IgG H&L 800CW at 1/20000 dilution

Lanes 1 - 6:

Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Observed band size: 40-60 kDa

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