Anti-TGF beta Receptor II antibody [EPR24349-124]

• IP

Supplier Data

Immunoprecipitation - Anti-TGF beta Receptor II antibody [EPR24349-124] (AB259360)

TGF beta Receptor II was immunoprecipitated from 0.35 mg HT-1080 (human fibrosarcoma epithelial cell) whole cell lysate 10 μg with ab259360 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab259360 at 1/1000 dilution. VeriBlot for IP secondary antibody (HRP) (ab131366) was used at 1/5000 dilution.

Lane 1 : HT-1080 (human fibrosarcoma epithelial cell) whole cell lysate 10 μg

Lane 2 : ab259360 IP in HT-1080 whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab259360 in HT-1080 whole cell lysate

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

Exposure time : 76 seconds.

Bands below 50 kDa are caused by degradation as demonstrated by WB data.

All lanes:

Immunoprecipitation - Anti-TGF beta Receptor II antibody [EPR24349-124] (ab259360)

Predicted band size: 65 kDa

Observed band size: 70-80 kDa

false

• IP

Supplier Data

Immunoprecipitation - Anti-TGF beta Receptor II antibody [EPR24349-124] (AB259360)

TGF beta Receptor II was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryonic fibroblast) whole cell lysate 10 μg with ab259360 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab259360 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate 10 μg

Lane 2 : ab259360 IP in NIH/3T3 whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab259360 in NIH/3T3 whole cell lysate

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

Exposure time : 50 seconds.

Bands below 50 kDa are caused by degradation as demonstrated by WB data.

All lanes:

Immunoprecipitation - Anti-TGF beta Receptor II antibody [EPR24349-124] (ab259360)

Predicted band size: 65 kDa

Observed band size: 70-80 kDa

false

• WB

Lab

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (AB259360)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. The molecular weight observed is consistent with what has been described in the literature (PMID : 23468914).

Lysates were made freshly and used in WB immediately to minimize protein degradation.

Exposure time : Lanes 1-2 : 81 seconds; Lane 3 : 3 minutes.

All lanes:

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (ab259360) at 1/1000 dilution

Lane 1:

HT-1080 (human fibrosarcoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

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

Predicted band size: 65 kDa

Observed band size: 70-80 kDa

false

• WB

Lab

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (AB259360)

Western blot : Rabbit Monoclonal [EPR24349-124] to TGF beta Receptor II ab259360 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta.

A band was observed at 70 kDa in Wild-type A549 cell lysates with no signal observed at this size in TGFBR2 knockout A549 cell line.

To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc Milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged.

Secondary antibodies used were Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (ab259360) at 1/1000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

Western blot - Human TGFBR2 (TGF beta Receptor II) knockout A549 cell line (<a href='/en-us/products/cell-lines/human-tgfbr2-tgf-beta-receptor-ii-knockout-a549-cell-line-ab277895'>ab277895</a>) at 20 µg

Lane 2:

TGFBR2 knockout A549 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 65 kDa

Observed band size: 70 kDa

false

• WB

Lab

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (AB259360)

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

Negative control : MCF7, PC-12 (PMID : 1310899).

Lysates used in this blot have experienced freeze-thaw cycles. Bands below 50 kDa are caused by degradation.

Exposure time : Lanes 1-4 : 81 seconds; Lane 5 : 3 minutes.

All lanes:

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (ab259360) at 1/1000 dilution

Lane 1:

HT-1080 (human fibrosarcoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

L6 (rat skeletal muscle myoblast) whole cell lysate at 20 µg

Lane 4:

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

Lane 5:

Rat lung 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

Predicted band size: 65 kDa

Observed band size: 70-80 kDa

false

• WB

Lab

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (AB259360)

Western blot : Anti-TGFBR2 antibody [EPR24349-124] (ab259360) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab259360 was shown to bind specifically to TGFBR2. A band was observed at 65 kDa in wild-type A549 cell lysates with no signal observed at this size in TGFBR2 knockout cell line. To generate this image, wild-type and TGFBR2 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (ab259360) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 40 µg

Lane 2:

TGFBR2 knockout A549 cell lysate at 40 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Observed band size: 65 kDa

false

• WB

Lab

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (AB259360)

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

Negative control : Neuro-2a (PMID : 1310899).

Lysates used in this blot have experienced freeze-thaw cycles. Bands below 50 kDa are caused by degradation.

Exposure time : 81 seconds.

All lanes:

Western blot - Anti-TGF beta Receptor II antibody [EPR24349-124] (ab259360) at 1/1000 dilution

Lane 1:

Mouse lung tissue lysate at 20 µg

Lane 2:

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

Lane 3:

Hepa1-6 (mouse hepatoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

Neuro-2a (mouse neuroblastoma neuroblast) 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: 65 kDa

Observed band size: 70-80 kDa

false