Anti-Bcl-2 抗体 [EPR17509]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Immunohistochemical analysis of paraffin-embedded Human endometrial cancer tissue labeling Bcl-2 with ab182858 at 1/1000 followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500.

Cytoplasm, nuclear membrane and nucleus staining on lymphocytes and cancer cells of Human endometrial cancer tissue is observed.

Counter stained with Hematoxylin.
Negative control : Used PBS instead of primary antibody followed by ab97051 at 1/500.

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-Bcl-2 antibody [EPR17509] (AB182858)

Immunohistochemical analysis of formalin fixed paraffin embedded human spleen labelling Bcl-2 with ab182858 at a concentration of 0.32µg/ml. The immunostaining was performed on a Leica Biosystems BOND ® RX instrument with a Bond™ Polymer Refine Detection kit. Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution 2) for 20mins.

ab182858 anti-Bcl-2 antibody [EPR17509] was incubated for 15mins at room temperature. Sections were counterstained with Hematoxylin. 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)

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Immunohistochemical analysis of formalin fixed paraffin embedded human spleen labelling Bcl-2 with ab182858 at a concentration of 0.86µg/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.

ab182858 anti-Bcl-2 antibody [EPR17509] 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

Supplier Data

Western blot - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Blocking and diluting buffer was 5% NFDM /TBST.

All lanes:

Western blot - Anti-Bcl-2 antibody [EPR17509] (ab182858) at 1/2000 dilution

Lane 1:

Human fetal kidney lysate at 10 µg

Lane 2:

Human fetal spleen lysate at 10 µg

Secondary

All lanes:

Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 26 kDa

Observed band size: 26 kDa

true

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Blocking and diluting buffer was 5% NFDM /TBST.

All lanes:

Western blot - Anti-Bcl-2 antibody [EPR17509] (ab182858) at 1/2000 dilution

Lane 1:

Mouse brain lysate at 10 µg

Lane 2:

Mouse heart lysate at 10 µg

Lane 3:

Mouse kidney lysate at 10 µg

Lane 4:

Mouse spleen lysate at 10 µg

Lane 5:

NIH/3T3 (Mouse embryo fibroblast cell line) whole cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/1000 dilution

Predicted band size: 26 kDa

Observed band size: 26 kDa

true

Exposure time: 3min

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed Jurkat (Human T cell leukemia cells from peripheral blood) cells labeling Bcl-2 with ab182858 at 1/250 (red) compared with a rabbit monoclonal IgG isotype control (ab172730) (black) and a unlabelled control (cells without incubation with primary antibody and secondary antibody (blue)). Goat anti rabbit IgG (FITC) at 1/500 was used as the secondary antibody.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling Bcl-2 with ab182858 at 1/1000 followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500.

Cytoplasm, nuclear membrane and nucleus staining on lymphocytes of Human tonsil tissue is observed.

Counter stained with Hematoxylin.

Negative control : Used PBS instead of primary antibody followed by ab97051 at 1/500.

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-Bcl-2 antibody [EPR17509] (AB182858)

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling Bcl-2 with ab182858 at 1/1000 followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500.

Cytoplasm, nuclear membrane and nucleus staining on lymphocytes of Mouse spleen tissue is observed.

Counter stained with Hematoxylin.
Negative control : Used PBS instead of primary antibody followed by ab97051 at 1/500.

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

• WB

Lab

Western blot - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Blocking/Diluting buffer 5% NFDM/TBST

All lanes:

Western blot - Anti-Bcl-2 antibody [EPR17509] (ab182858) at 1/10000 dilution

Lane 1:

NIH/3T3 (mouse embryo fibroblast cell line) whole cell lysate at 20 µg

Lane 2:

WEHI-3 (mouse leukemia cell line) whole cell lysate at 20 µg

Lane 3:

Mouse hippocampus at 10 µg

Lane 4:

Mouse heart at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG H&L (HRP) at 1/2000 dilution

Predicted band size: 26 kDa

Observed band size: 26 kDa

false

Exposure time: 8s

• WB

Lab

Western blot - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Lanes 1- 2 : Merged signal (red and green). Green - ab182858 observed at 26 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

ab182858 was shown to react with Bcl-2 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255364 (knockout cell lysate ab263752) was used. Wild-type HeLa and BCL2 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab182858 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 2000 dilution and a 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-Bcl-2 antibody [EPR17509] (ab182858) at 1/2000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

BCL2 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human BCL2 knockout HeLa cell line (<a href='/products/cell-lines/human-bcl2-knockout-hela-cell-line-ab255364'>ab255364</a>)

Predicted band size: 26 kDa

Observed band size: 26 kDa

false

• WB

Supplier Data

Western blot - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Blocking and diluting buffer was 5% NFDM /TBST.

All lanes:

Western blot - Anti-Bcl-2 antibody [EPR17509] (ab182858) at 1/20000 dilution

Lane 1:

Human tonsil lysate at 20 µg

Lane 2:

Human thymus lysate at 20 µg

Lane 3:

Jurkat (Human T cell leukemia cells from peripheral blood) whole cell lysate at 20 µg

Lane 4:

U-937 (Human histiocytic lymphoma cells) whole cell lysate at 20 µg

Lane 5:

THP-1 (Human monocytic leukemia cells) whole cell lysate at 20 µg

Lane 6:

HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 7:

C2C12 (Mouse myoblast cell line) whole cell lysate at 20 µg

Lane 8:

WEHI-3 (Mouse leukemia cell line) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/1000 dilution

Predicted band size: 26 kDa

Observed band size: 26 kDa

true

Exposure time: 1min

• WB

Lab

Western blot - Anti-Bcl-2 antibody [EPR17509] (AB182858)

Lanes 1 - 4 : Merged signal (red and green). Green - ab182858 observed at 26 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab182858 was shown to specifically react with BCL2 when BCL2 knockout samples were used. Wild-type and BCL2 knockout samples were subjected to SDS-PAGE. ab182858 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1 ug/ml and 1/10000 dilution respectively. Blots were developed 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 1/10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Bcl-2 antibody [EPR17509] (ab182858) at 1 µg/mL

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

BCL2 knockout HAP1 whole cell lysate at 20 µg

Lane 3:

HeLa whole cell lysate at 20 µg

Lane 4:

THP-1 whole cell lysate at 20 µg

Predicted band size: 26 kDa

Observed band size: 26 kDa

false