Anti-TGF beta 3 antibody [EPR27093-72]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized PC-12 (rat adrenal gland pheochromocytoma small irregularly shaped cells) cells labelling TGF beta 3 with ab308300 at 1/250 (1.808 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2ug/mL dilution (Green). Confocal image showing cytoplasmic staining in PC-12 cell line.Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2ug/mL dilution.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized A549 (human lung carcinoma epithelial cell) cells labelling TGF beta 3 with ab308300 at 1/250 (1.808 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2ug/mL dilution (Green). Confocal image showing cytoplasmic staining in A549 cell line.Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Red). The Nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2ug/mL dilution.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized A549 (human lung carcinoma epithelial cell) cells labelling TGF beta 3 with ab308300 at 1/500 dilution (0.1 ug)/Red (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• IP

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Immunoprecipitation - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

TGF beta 3 was immunoprecipitated from 0.35 mg A549 (human lung carcinoma epithelial cell) whole cell lysate with ab308300 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab308300 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : A549 (human lung carcinoma epithelial cell) whole cell lysate Lane 2 : abAB308300 IP in A549 (human lung carcinoma epithelial cell) whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab308300 in A549 whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 24 seconds

All lanes:

Immunoprecipitation - Anti-TGF beta 3 antibody [EPR27093-72] (ab308300) at 1/30 dilution

All lanes:

A549 (human lung carcinoma epithelial cell) whole cell lysate at 4 µg

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 55 kDa

false

Exposure time: 24s

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized PC-12 (rat adrenal gland pheochromocytoma cell) cells labelling TGF beta 3 with ab308300 at 1/500 dilution (0.1 ug)/Red (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• WB

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Western blot - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. Lysates were freshly made and used for Western blotting immediately to minimize protein degradation. Precursor/latent complex observed at 55kDa. Exposure time : 81 seconds

All lanes:

Western blot - Anti-TGF beta 3 antibody [EPR27093-72] (ab308300) at 1/1000 dilution

Lane 1:

HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

PC-12 (rat adrenal gland pheochromocytoma cell) 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

Predicted band size: 47 kDa

Observed band size: 55 kDa

false

Exposure time: 81s

• WB

Supplier Data

Western blot - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. The expression of TGF beta 3 is upregulated in response to CoCl2 treatment (PMID : 29577601). In Western blot, anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution. Precursor/latent complex observed at 55kDa. Exposure time : 15 seconds.

All lanes:

Western blot - Anti-TGF beta 3 antibody [EPR27093-72] (ab308300) at 1/1000 dilution

Lane 1:

Untreated Saos-2 (human osteosarcoma epithelial) whole cell lysate, at 20 µg

Lane 2:

Saos-2 treated with 250uM CoCl2 for 24h 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

Predicted band size: 47 kDa

Observed band size: 55 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. Lysates were freshly made and used for Western blotting immediately to minimize protein degradation. Exposure time : 15 seconds.

All lanes:

Western blot - Anti-TGF beta 3 antibody [EPR27093-72] (ab308300) at 1/1000 dilution

All lanes:

A549 (human lung carcinoma epithelial cell) 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

Predicted band size: 47 kDa

Observed band size: 55 kDa

false

Exposure time: 15s

• Dot

Supplier Data

Dot Blot - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Dot blot analysis of TGF beta 3 using ab308300 at 1 : 1000 (0.452 ug/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution. Exposure time : 180 seconds Blocking and diluting buffer and concentration : 5% NFDM/TBST

• WB

Supplier Data

Western blot - Anti-TGF beta 3 antibody [EPR27093-72] (AB308300)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. Precursor/latent complex observed at 55kDa. Exposure time : 180 seconds.

All lanes:

Western blot - Anti-TGF beta 3 antibody [EPR27093-72] (ab308300) at 1/1000 dilution

Lane 1:

Mouse kidney tissue lysate at 80 µg

Lane 2:

Mouse liver tissue lysate, at 80 µg

Lane 3:

Mouse lung tissue lysate at 80 µg

Lane 4:

Mouse colon tissue lysate at 80 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

Predicted band size: 47 kDa

Observed band size: 55 kDa

false

Exposure time: 180s