Anti-Topoisomerase I antibody [EPR5375]

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Immunocytochemistry/ Immunofluorescence analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling Topoisomerase I with Purified ab109374 at 1/500 dilution (0.3 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 nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Immunohistochemical analysis of paraffin-embedded Human breast carcinoma tissue using ab109374 at a dilution of 1/100. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human clear cell carcinoma of kidney tissue sections labeling Topoisomerase I with Purified ab109374 at 1 : 100 dilution (1.29 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Intracellular Flow Cytometry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling Topoisomerase I with Purified ab109374 at 1/20 dilution (10μg/ml) (red). Cells were fixed with 80% Methanol. A Goat anti rabbit IgG (Alexa Fluorr^® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Overlay histogram showing HepG2 cells stained with ab109374 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab109374, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluorr^® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1μg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Immunohistochemical analysis of paraffin-embedded Human colonic carcinoma tissue using ab109374 at a dilution of 1/100. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Immunofluorescent staining of MCF7 cells using ab109374 at a dilution of 1/100.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Rat colon tissue sections labeling Topoisomerase I with purified ab109374 at 1 : 100 dilution (1.29 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Mouse kidney tissue sections labeling Topoisomerase I with purified ab109374 at 1 : 100 dilution (1.29 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• WB

Lab

Western blot - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

All lanes:

Western blot - Anti-Topoisomerase I antibody [EPR5375] (ab109374) at 1/10000 dilution

Lane 1:

MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates at 20 µg

Lane 2:

Jurkat (Human T cell leukemia T lymphocyte) whole cell lysates at 20 µg

Lane 3:

HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysates at 20 µg

Lane 4:

Neuro-2a (Mouse neuroblastoma neuroblast) whole cell lysates 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 1/20000 dilution

Predicted band size: 91 kDa

Observed band size: 91 kDa

false

• WB

Lab

Western blot - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

This blot was produced using 4-20% SDS-PAGE containing 15 μg of Jurkat whole cell lysate per lane at 150V for 1hr before being transferred onto a 0.45 μm PVDF membrane at 75V for 1hr. The membrane was then blocked for 1hr using 5% NFDM/TBST, then incubated with ab109374 (1/10,000) at room temperature for 1hr. After being washed three times in TBST, the membrane was incubated with Peroxidase conjugated goat anti-rabbit IgG (H+L) (ab97051) at 1/20,000 dilution for 1hr at room temperature. The membrane was washed three times again. Then the signal was developed using the ECL technique. ab109374 was stored at a range of temperatures (+4°C, +22°C, +37°C) for 1 week before being tested in WB. The image shows the band intensity remains relatively constant across all storage temperatures, demonstrating that antibody activity is not affected.

All lanes:

Western blot - Anti-Topoisomerase I antibody [EPR5375] (ab109374) at 1/10000 dilution

All lanes:

Jurkat whole cell lysate at 15 µg with NDFM/TBST

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

false

Exposure time: 60s

• WB

Lab

Western blot - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

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

All lanes:

Western blot - Anti-Topoisomerase I antibody [EPR5375] (ab109374) at 1/50000 dilution

All lanes:

PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysates 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 1/20000 dilution

Predicted band size: 91 kDa

Observed band size: 91 kDa

false

• WB

Unknown

Western blot - Anti-Topoisomerase I antibody [EPR5375] (AB109374)

All lanes:

Western blot - Anti-Topoisomerase I antibody [EPR5375] (ab109374) at 1/10000 dilution

Lane 1:

Jurkat cell lysate at 10 µg

Lane 2:

MCF7 cell lysate at 10 µg

Lane 3:

SW480 cell lysate at 10 µg

Lane 4:

K562 cell lysate at 10 µg

Predicted band size: 91 kDa

false