Anti-TLE 1 antibody [EPR9386(2)]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Immunofluorescence analysis of 4% paraformaldehyde-fixed, 0.1% TritonX-100 permeabilised wildtype HEK293T cells and TLE1 knockout HEK293T cells (ab265059) with ab183742 (green) at 1/50 dilution. Alexa Fluor® 488 Goat Anti-Rabbit IgG H&L (ab150081) was used as a secondary antibody, presabsorbed at 1/1000 dilution. Alexa Fluor® 594 Anti-alpha Tubulin mouse monoclonal antibody (ab195889) used as microtubule marker counterstain (red). Nuclei were counterstained with DAPI (blue). Confocal image showing nuclear staining in wildtype HEK293T cells and showing no staining in TLE1 knockout HEK293T cells. Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Immunofluorescence analysis of paraformaldehyde-fixed HepG2 cells staining TLE 1 (green) with ab183742 at 1/100 dilution. Alexa Fluor®488-conjugated goat anti rabbit IgG was used as a secondary antibody at 1/200 dilution. Nuclei were counterstained with DAPI (blue).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

ab183742 staining TLE1 in wild-type MCF7 cells (top panel) and TLE1 knockout MCF7 cells (bottom panel). The cells were fixed with 4% paraformaldehyde (10 min), 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 ab183742 at 1/500 dilution 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 high-content analysis system (Perkin Elmer, Operetta CLS™).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Immunohistochemical analysis of Human schwannoma, staining TLE 1 with ab183742 at 1/250 dilution. Detected using HRP Polymer for Rabbit IgG and counter-stained using hematoxylin.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Immunohistochemical analysis of Human synovial sarcoma, staining TLE 1 with ab183742 at 1/250 dilution. Detected using HRP Polymer for Rabbit IgG and counter-stained using hematoxylin.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Immunohistochemical analysis of paraffin-embedded fixed (A) Parental HEK293 (Human embryonic kidney epithelial cell) cell pellet (B) TLE1 knockout HEK293 (ab265059) cell pellet, staining TLE 1 with ab183742 at 1/250 dilution for 30 mins at room temperature. LeicaDS9800 (Bond™ Polymer Refine Detection) used as secondary antibody. Counter-stained using hematoxylin. Positive staining on Wild-type HEK293T cell pellet and no staining on TLE1 knockout HEK293 cell pellet.  The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.

• WB

Lab

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Lanes 1-3 : Merged signal (red and green). Green - ab183742 observed at 83 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab183742 Anti-TLE 1 antibody [EPR9386(2)] was shown to specifically react with TLE 1 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab265059 (knockout cell lysate ab257240) was used. Wild-type and TLE 1 knockout samples were subjected to SDS-PAGE. ab183742 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C 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 10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (ab183742) at 1/1000 dilution

Lane 1:

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

Lane 2:

TLE1 knockout HEK-293T cell lysate at 20 µg

Lane 3:

HepG2 cell lysate at 20 µg

Predicted band size: 83 kDa

Observed band size: 83 kDa

false

• WB

Supplier Data

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

All lanes:

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (ab183742) at 1/1000 dilution

Lane 1:

SH-SY5Y cell lysate at 20 µg

Lane 2:

HepG2 cell lysate at 20 µg

Lane 3:

Jurkat cell lysate at 20 µg

Lane 4:

HeLa cell lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 83 kDa

Observed band size: 83 kDa

false

• WB

Lab

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Lanes 1 - 4 : Merged signal (red and green). Green - ab183742 observed at 83 kDa. Red - loading control, ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.

ab183742 was shown to react with TLE 1 in western blot. The band observed in the CRISPR/Cas9 edited lysate lane below 83 kDa is likely to represent a truncated form. This has not been investigated further. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab183742 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (ab183742) at 1/1000 dilution

Lane 1:

Wild-type MCF7 cell lysate at 20 µg

Lane 2:

TLE1 CRISPR/Cas9 edited MCF7 cell lysate at 20 µg

Lane 2:

Western blot - Human TLE1 knockout MCF7 cell line (<a href='/en-us/products/cell-lines/human-tle1-knockout-mcf7-cell-line-ab269498'>ab269498</a>)

Lane 3:

SH-SY5Y cell lysate at 20 µg

Lane 4:

HepG2 cell lysate at 20 µg

Predicted band size: 83 kDa

Observed band size: 83 kDa

false

• WB

Unknown

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Lanes 1-3 : Merged signal (red and green). Green - ab183742 observed at 83 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab183742 Anti-TLE 1 antibody [EPR9386(2)] was shown to specifically react with TLE 1 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab265059 (knockout cell lysate ab257240) was used. Wild-type and TLE 1 knockout samples were subjected to SDS-PAGE. ab183742 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C 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 10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (ab183742) at 1/1000 dilution

Lane 1:

Wild-type HEK293T cell lysate at 20 µg

Lane 2:

Western blot - Human TLE1 knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-tle1-knockout-hek-293t-cell-line-ab265059'>ab265059</a>) at 20 µg

Lane 2:

TLE1 knockout HEK293T cell lysate at 20 µg

Lane 3:

HepG2 cell lysate at 20 µg

Predicted band size: 83 kDa

Observed band size: 83 kDa

false

• WB

Lab

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Lanes 1-3 : Merged signal (red and green). Green - ab183742 observed at 83 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab183742 Anti-TLE 1 antibody [EPR9386(2)] was shown to specifically react with TLE 1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264901 (knockout cell lysate ab257241) was used. Wild-type and TLE 1 knockout samples were subjected to SDS-PAGE. ab183742 and Anti-GAPDH antibody [EPR16891] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 500 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 10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (ab183742) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

TLE1 knockout HeLa cell lysate at 20 µg

Lane 3:

293T cell lysate at 20 µg

Predicted band size: 83 kDa

Observed band size: 83 kDa

false

• WB

Lab

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Lanes 1-3 : Merged signal (red and green). Green - ab183742 observed at 83 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab183742 Anti-TLE 1 antibody [EPR9386(2)] was shown to specifically react with TLE 1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264901 (knockout cell lysate ab257241) was used. Wild-type and TLE 1 knockout samples were subjected to SDS-PAGE. ab183742 and Anti-GAPDH antibody [EPR16891] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 500 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 10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (ab183742) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

TLE1 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human TLE1 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-tle1-knockout-hela-cell-line-ab264901'>ab264901</a>)

Lane 3:

293T cell lysate at 20 µg

Predicted band size: 83 kDa

Observed band size: 83 kDa

false

• WB

Lab

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (AB183742)

Lanes 1 - 4 : Merged signal (red and green). Green - ab183742 observed at 83 kDa. Red - loading control, ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.

ab183742 was shown to react with TLE 1 in western blot. The band observed in the knockout cell line ab269498 (knockout cell lysate ab269660) lane below 83 kDa is likely to represent a truncated form. This has not been investigated further. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab183742 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-TLE 1 antibody [EPR9386(2)] (ab183742) at 1/1000 dilution

Lane 1:

Wild-type MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Lane 2:

TLE1 knockout MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Lane 3:

SH-SY5Y (Human neuroblastoma cell line from bone marrow) whole cell lysate at 20 µg

Lane 4:

Hep G2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg

Predicted band size: 83 kDa

Observed band size: 83 kDa

false