Anti-Smac/Diablo antibody [Y12]

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (AB32023)

Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling Smac / Diablo with ab32023 at 1/500. Cells were fixed with 100% methanol. ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody.
Control : PBS only.
Nuclear counter stain : DAPI.

• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (AB32023)

Immunofluorescence analysis of IMR32 cells, staining Smac / Diablo with ab32023. Cells were fixed with 4% paraformaldehyde before incubating with primary antibody. An Alexa Fluor^® conjugated anti-rabbit IgG was used as the secondary antibody.

Image from Lamers F et al. BMC Cancer. 2012 Jul 12;12:285. doi: 10.1186/1471-2407-12-285. Fig 3.; 12 July 2012 BMC Cancer 2012 12:285.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smac/Diablo antibody [Y12] (AB32023)

ab32023, at a 1/10 dilution, staining human Smac/Diablo in skin cancer tissue by immunohistochemistry, paraffin-embedded tissue.

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

• IP

Lab

Immunoprecipitation - Anti-Smac/Diablo antibody [Y12] (AB32023)

Smac/Diablo was immunoprecipitated from 0.35 mg Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 μg with ab32023 at 1/50 dilution (2μg). VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/5000 dilution.

Lane 1 : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 μg

Lane 2 : ab32023 IP in Jurkat whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab32023 in Jurkat whole cell lysate

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

All lanes:

Immunoprecipitation - Anti-Smac/Diablo antibody [Y12] (ab32023)

Predicted band size: 27 kDa

Observed band size: 21 kDa

false

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (AB32023)

Immunocytochemistry/Immunofluorescence analysis of NIH/3T3 (mouse embryonic fibroblast) cells labelling Smac / Diablo with ab32023 at 1/100 dilution (green). Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100. ab150081, a Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (1/1000) was used as the secondary antibody. Anti-COX IV mouse monoclonal antibody - Mitochondrial Marker at 1/1000 was as the counterstain antibody followed by ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 (magenta). Nuclear DNA was labeled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). Negative Control 1 : ab32023 at 1/100 followed by ab150120 at 1/1000. Negative Control 2 : anti-COX IV antibody at 1/1000 followed by ab150081 at 1/1000. Confocal image showing positive staining in mitochondria of NIH/3T3 cell line (shown in green).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smac/Diablo antibody [Y12] (AB32023)

Immunohistochemistry analysis of paraffin-embedded mouse brain tissue sections labeling Smac/Diablo with ab32023 at 1/6000 dilution, incubated for 30 mins at room temperature followed by ab209101 (ready to use). Sections were counterstained with hematoxylin. Antigen retrieval was heat mediated with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Positive staining on mouse brain.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smac/Diablo antibody [Y12] (AB32023)

Immunohistochemistry analysis of paraffin-embedded mouse testis tissue sections labeling Smac/Diablo with ab32023 at 1/6000 dilution, incubated for 30 mins at room temperature followed by ab209101 (ready to use). Sections were counterstained with hematoxylin. Antigen retrieval was heat mediated with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Smac/Diablo antibody [Y12] (AB32023)

Immunocytochemistry/Immunofluorescence analysis of C6 (rat glial tumor glial cell) cells labelling Smac / Diablo with ab32023 at 1/100 dilution (green). Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100. ab150081, a Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (1/1000) was used as the secondary antibody. Anti-COX IV mouse monoclonal antibody - Mitochondrial Marker at 1/1000 was as the counterstain antibody followed by ab150120, Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 (magenta). Nuclear DNA was labeled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). Negative Control 1 : ab32023 at 1/100 followed by ab150120 at 1/1000. Negative Control 2 : anti-COX IV antibody at 1/1000 followed by ab150081 at 1/1000. Confocal image showing positive staining in mitochondria of C6 cell line (shown in green).

• WB

Unknown

Western blot - Anti-Smac/Diablo antibody [Y12] (AB32023)

All lanes:

Western blot - Anti-Smac/Diablo antibody [Y12] (ab32023) at 1/1000 dilution

All lanes:

Jurkat cell lysate

Predicted band size: 27 kDa

Observed band size: 21 kDa

false

• WB

Lab

Western blot - Anti-Smac/Diablo antibody [Y12] (AB32023)

Loading control : Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1/200000) (36KDa). Blocking/Diluting buffer and concentration 5% NFDM/TBST Low expression : spleen, brain, heart (PMID : 12058276)

Lanes 1 - 8:

Western blot - Anti-Smac/Diablo antibody [Y12] (ab32023) at 1/1000 dilution

Lanes 1 - 8:

Western blot - Anti-Smac/Diablo antibody [Y12] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/smac-diablo-antibody-y12-bsa-and-azide-free-ab239797'>ab239797</a>)

Lane 1:

Mouse testis tissue lysate at 20 µg

Lane 2:

Mouse brain tissue lysate at 20 µg

Lane 3:

Mouse heart tissue lysate at 20 µg

Lane 4:

Mouse spleen tissue lysate at 20 µg

Lane 5:

Rat testis tissue lysate at 20 µg

Lane 6:

Rat brain tissue lysate at 20 µg

Lane 7:

Rat heart tissue lysate at 20 µg

Lane 8:

Rat spleen 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

false

Exposure time: 10s

• WB

Lab

Western blot - Anti-Smac/Diablo antibody [Y12] (AB32023)

Loading control : Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1/200000) (36KDa). Blocking/Diluting buffer and concentration 5% NFDM/TBST

Lanes 1 - 4:

Western blot - Anti-Smac/Diablo antibody [Y12] (ab32023) at 1/1000 dilution

Lanes 1 - 4:

Western blot - Anti-Smac/Diablo antibody [Y12] - BSA and Azide free (<a href='/en-us/products/primary-antibodies/smac-diablo-antibody-y12-bsa-and-azide-free-ab239797'>ab239797</a>)

Lane 1:

C6 (rat glial tumor glial cell) whole cell lysate at 20 µg

Lane 2:

RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 3:

PC-12 (rat adrenal gland pheochromocytoma cell) whole cell lysate at 20 µg

Lane 4:

NIH/3T3 (mouse embryonic fibroblast) whole cell 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

false

Exposure time: 37s