Anti-SOAT 1/ACAT1 antibody [EPR26952-40]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (AB307597)

Immunohistochemical analysis of paraffin-embedded human cerebellum tissue labeling SOAT 1/ACAT1 with ab307597 at 1/1000 dilution (0.47 µg/ml) followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Negative control : no staining on human cerebellum. The section was incubated with ab307597 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (AB307597)

Immunohistochemical analysis of paraffin-embedded human testis tissue labeling SOAT 1/ACAT1 with ab307597 at 1/1000 dilution (0.47 µg/ml) followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on Leydig cells of the human testis (PMID : 10623671). The section was incubated with ab307597 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (AB307597)

Immunohistochemical analysis of paraffin-embedded human adrenal gland tissue labeling SOAT 1/ACAT1 with ab307597 at 1/1000 dilution (0.47 µg/ml) followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on cortex of human adrenal gland (PMID : 31265901). The section was incubated with ab307597 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (AB307597)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labeling SOAT 1/ACAT1 with ab307597 at 1/250 dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2µg/ml) (Green). Confocal image showing the cytoplasmic staining co-located with endoplasmic reticulum marker in parental Hela cell line, but no staining in SOAT1 KO HeLa cell line. Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8). Anti-KDEL mouse monoclonal antibody was used to counterstain at 1/200 dilution (5 µg/ml) followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 dilution (2µg/ml) (Red). Nuclear counterstain was DAPI (Blue). -ve control 1 : ab307597 at 1/250 dilution followed by ab150120 at a 1/1000 dilution. -ve control 2 : Anti-KDEL mouse monoclonal antibody at 1/200 dilution followed by ab150081 at a 1/1000 dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (AB307597)

Immunohistochemical analysis of paraffin-embedded cell pellets labeling SOAT 1/ACAT1 with ab307597 at 1/1000 dilution (0.47 µg/ml) followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on (A) wild-type HeLa cell pellet, no staining on (B) SOAT1 knockout HeLa cell pellet. The section was incubated with ab307597 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins.

• IP

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Immunoprecipitation - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (AB307597)

SOAT 1/ACAT1 was immunoprecipitated from 0.35 mg HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate 10 µg with ab307597 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307597 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : HepG2 whole cell lysate 10 µg Lane 2 : ab307597 IP in HepG2 whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab307597 in HepG2 whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 5 seconds. The molecular weight observed is consistent with what has been described in the literature (PMID : 25617738; 32424158). The bands above 50kDa are dimer and tetramer.

All lanes:

Immunoprecipitation - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (ab307597) at 1/30 dilution

All lanes:

HepG2 (human hepatocellular carcinoma epithelial cell)

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 50 kDa,100 kDa,150 kDa

false

Exposure time: 5s

• WB

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Western blot - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (AB307597)

Blocking and diluting buffer and concentration : Ѕ volume of Odyssey Blocking Buffer (TBS)+ Ѕ volume of 0.1% TBS. The samples were run on a Bis-Tris gel. Performed under reducing conditions. False colour image of Western blot : Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (ab307597) 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, ab307597 was shown to bind specifically to SOAT 1/ACAT1. A band was observed at 50/100 kDa in wild-type HeLa cell lysates whereas no signal observed at this size in SOAT 1/ACAT1 knockout cell line ab264862 (knockout cell lysate ab257695). To generate this image, wild-type and SOAT 1/ACAT1 knockout HeLa cell lysates were analyzed. First, samples were run on a SDS-PAGE gel then transferred onto an immobilon-FL PVDF membrane. Membranes were blocked in in Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS 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) at 1/10000 dilution and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution. Low expression : HT-29 (PMID : 35821230).

All lanes:

Western blot - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (ab307597) at 1/1000 dilution

Lane 1:

Wild-type HeLa whole cell lysate at 20 µg

Lane 2:

SOAT 1/ACAT1 knockout HeLa whole cell lysate at 20 µg

Lane 3:

HepG2 (human hepatocellar carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

HT-29 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Secondary

Lanes 1 - 4:

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

Lanes 1 - 4:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/10000 dilution

Observed band size: 50 kDa,100 kDa,150 kDa

false

• WB

Supplier Data

Western blot - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (AB307597)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. The molecular weight observed is consistent with what has been described in the literature (PMID : 25617738; 32424158). The bands above 50kDa are dimer and tetramer. Negative control : cerebellum. Exposure times : Lanes 1-2 : 26 seconds Lanes 3-5 : 81 seconds

All lanes:

Western blot - Anti-SOAT 1/ACAT1 antibody [EPR26952-40] (ab307597) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HepG2 (human hepatocellar carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

Human testis tissue lysate at 20 µg

Lane 4:

Human colon tissue lysate at 20 µg

Lane 5:

Human cerebellum tissue lysate at 20 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 1/100000 dilution

Observed band size: 50 kDa,100 kDa,150 kDa

false