Anti-ACSS2 antibody [EPR8500]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-ACSS2 antibody [EPR8500] (AB133664)

Immunofluorescence analysis of ACSS2 in Caco 2 cells labelled with unpurified ab133664 at a 1/100 dilution.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-ACSS2 antibody [EPR8500] (AB133664)

Immunocytochemistry/ Immunofluorescence analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling ACSS2 with purified ab133664 at 1 : 200 dilution (8.5 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.

• WB

Lab

Western blot - Anti-ACSS2 antibody [EPR8500] (AB133664)

Western blot : Rabbit Monoclonal[EPR8500] to ACSS2 ab133664 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 75 kDa in Wild-type U-87 MG ab278079 cell lysates with no signal observed at this size in ACSS2 knockout U-87 MG cell line (ab326072). To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-ACSS2 antibody [EPR8500] (ab133664) at 1/1000 dilution

Lane 1:

Wild-type U-87 MG ab278079 at 20 µg

Lane 2:

Western blot - Human ACSS2 knockout U-87 MG cell line (<a href='/en-us/products/cell-lines/human-acss2-knockout-u-87-mg-cell-line-ab326072'>ab326072</a>) at 20 µg

Lane 3:

Wild-type HAP1 at 20 µg

Lane 4:

ACSS2 knockout HAP1 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 79 kDa

Observed band size: 75 kDa

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• WB

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Western blot - Anti-ACSS2 antibody [EPR8500] (AB133664)

All lanes:

Western blot - Anti-ACSS2 antibody [EPR8500] (ab133664) at 1/1000 dilution

Lane 1:

U87 MG lysate at 10 µg

Lane 2:

HepG2 lysate at 10 µg

Lane 3:

Caco 2 lysate at 10 µg

Lane 4:

Fetal liver lysate at 10 µg

Predicted band size: 79 kDa

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• WB

Lab

Western blot - Anti-ACSS2 antibody [EPR8500] (AB133664)

All lanes:

Western blot - Anti-ACSS2 antibody [EPR8500] (ab133664) at 1/1000 dilution

Lane 1:

U-87 MG (Human glioblastoma-astrocytoma epithelial cell) whole cell lysates at 15 µg

Lane 2:

Mouse brain lysates at 15 µg

Lane 3:

Rat brain lysates at 15 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 79 kDa

Observed band size: 78 kDa

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• WB

Lab

Western blot - Anti-ACSS2 antibody [EPR8500] (AB133664)

False colour image of Western blot : Anti-ACSS2 antibody [EPR8500] 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, ab133664 was shown to bind specifically to ACSS2. A band was observed at 75 kDa in wild-type HAP1 cell lysates with no signal observed at this size in ACSS2 knockout cell line HAP1. To generate this image, wild-type and ACSS2 knockout HAP1 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-ACSS2 antibody [EPR8500] (ab133664) at 1/1000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 40 µg

Lane 2:

ACSS2 knockout HAP1 cell lysate at 40 µg

Lane 3:

HepG2 cell lysate at 20 µg

Lane 4:

U-87 MG cell lysate at 20 µg

Predicted band size: 79 kDa

Observed band size: 75 kDa

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• WB

CiteAb

Western blot - Anti-ACSS2 antibody [EPR8500] (AB133664)

ACSS2 western blot using anti-ACSS2 antibody [EPR8500] ab133664. Publication image and figure legend from He, W., Liang, B., et al., 2019, Oncogene, PubMed 30742067.

ab133664 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab133664 please see the product overview.

Fatty acid oxidation plays an important role in mesenchymal stem cell (MSC)-induced stemness and chemoresistance. a Expression levels of carnitine palmitoyltransferase 1 (CPT1) and acetyl-coenzyme A synthetase (ACS) in AGS and MKN45 co-culture with MSCs compared to culture alone by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. b qRT-PCR for CPT1 and ACS in AGS and MKN45 sphere culture either alone or with MSCs. c–e CPT1 enzyme activity (c), relative fatty β-oxidation rate (d), and ATP levels (e) in AGS and MKN45 cells with or without MSCs. f Expression levels of stemness-associating genes in AGS and MKN45 transfected with siCPT1. g, h qRT-PCR (g) and western blot (h) for stemness-associating genes in AGS and MKN45 cell culture either alone or with MSCs and with or without 100 μmol/L etomoxir (ETX). i Representative images of sphere-formation assay in AGS and MKN45 cell culture either alone or with MSCs and with or without 100 μmol/L ETX for 7 days. Scale bar = 500 μm. j Colony-formation assay and the quantitative graph of AGS and MKN45 cell culture either alone or with MSCs and with or without 100 μmol/L ETX when treated with 1 μg/mL 5-florouracil and 3 μg/mL oxaliplatin. k ATP level in AGS and MKN45 cell culture either alone or with MSCs and with or without 100 μmol/L ETX for 48 h. *p < 0.05; **p < 0.01; ***p < 0.001

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