Anti-SOX17 antibody [EPR20684]

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-SOX17 antibody [EPR20684] (AB224637)

ab224637 staining SOX17 in wild-type HeLa cells (top panel) and SOX17 knockout HeLa cells (ab265744) (bottom panel). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab224637 at 0.2μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor^® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor^® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOX17 antibody [EPR20684] (AB224637)

Immunohistochemical analysis of paraffin-embedded human choriocarcinoma tissue labeling SOX17 with ab224637 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Negative tissue : no staining on tumor cells of human choriocarcinoma (PMID : 19369635) is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOX17 antibody [EPR20684] (AB224637)

Immunohistochemical analysis of formalin fixed paraffin embedded human spleen labelling SOX17 with ab224637 at a concentration of 0.5µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with a OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was performed with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5 for 32mins. ab224637 anti-SOX17 antibody [EPR20684] was incubated for 16mins at 37°C. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual).

• WB

Lab

Western blot - Anti-SOX17 antibody [EPR20684] (AB224637)

Exposure time :
Lanes 1-2 : 5 seconds
Lanes 3-6 : 3 minutes

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression profile observed is consistent with the literature (PMID : 11786926).

Negative control : HeLa.

All lanes:

Western blot - Anti-SOX17 antibody [EPR20684] (ab224637) at 1/500 dilution

Lane 1:

SK-OV-3 (Human adenocarcinoma) whole cell lysates at 20 µg

Lane 2:

NIH: OVCAR-3 (Human ovary adenocarcinoma) whole cell lysates at 20 µg

Lane 3:

HeLa (Human cervix adenocarcinoma) whole cell lysates at 20 µg

Lane 4:

Rat E14 embryo tissue lysates at 20 µg

Lane 5:

Rat E9.5 embryo tissue lysates at 20 µg

Lane 6:

Rat lung tissue lysates at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG) at 1/2000 dilution

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• WB

Lab

Western blot - Anti-SOX17 antibody [EPR20684] (AB224637)

Lanes 1-3 : Merged signal (red and green). Green - ab224637 observed at 51 kDa. Red - loading control ab8245 observed at 36 kDa.

ab224637 Anti-SOX17 antibody [EPR20684] was shown to specifically react with SOX17 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265744 (knockout cell lysate ab257697) was used. Wild-type and SOX17 knockout samples were subjected to SDS-PAGE. ab224637 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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-SOX17 antibody [EPR20684] (ab224637) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

SOX17 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human SOX17 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-sox17-knockout-hela-cell-line-ab265744'>ab265744</a>)

Lane 3:

SK-OV-3 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: 44 kDa

Observed band size: 51 kDa

false

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOX17 antibody [EPR20684] (AB224637)

Immunohistochemical analysis of paraffin-embedded rat lung tissue labeling SOX17 with ab224637 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Nuclear staining on endothelium of rat lung (PMID : 24418654) is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOX17 antibody [EPR20684] (AB224637)

Immunohistochemical analysis of paraffin-embedded mouse spleen tissue labeling SOX17 with ab224637 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Nuclear staining on endothelium of mouse spleen (PMID : 24418654) is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SOX17 antibody [EPR20684] (AB224637)

Immunohistochemical analysis of paraffin-embedded human seminoma tissue labeling SOX17 with ab224637 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Nuclear staining on tumor cells of human seminoma (PMID : 19369635; PMID : 18348160) is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IP

Supplier Data

Immunoprecipitation - Anti-SOX17 antibody [EPR20684] (AB224637)

SOX17 was immunoprecipitated from 0.35 mg of SK-OV-3 (human ovarian cancer cell line) lysate with ab224637 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab224637 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

Lane 1 : SK-OV-3 whole cell lysate 10 μg (Input).

Lane 2 : ab224637 IP in SK-OV-3 whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab224637 in SK-OV-3 whole cell lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 1 second.

All lanes:

Immunoprecipitation - Anti-SOX17 antibody [EPR20684] (ab224637)

Predicted band size: 44 kDa

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