Anti-TYK2 antibody [EPR24628-106]

• IP

Lab

Immunoprecipitation - Anti-TYK2 antibody [EPR24628-106] (AB303500)

TYK2 was immunoprecipitated from 0.35 mg HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate with ab303500 at 1/30 dilution (2 µg in 0.35 mg lysates). Western blot was performed on the immunoprecipitate using ab303500 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP) (ab131366) was used at 1/5000 dilution.

All lanes:

Immunoprecipitation - Anti-TYK2 antibody [EPR24628-106] (ab303500) at 1/1000 dilution

Lane 1:

Hela (human cervical adenocarcinoma epithelial cell) whole cell lysate at 10 µg

Lane 2:

ab303500 at 1/30 IP in Hela (human cervical adenocarcinoma epithelial cell) whole cell lysate at 10 µg

Lane 3:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab303500 in Hela whole cell lysate at 10 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

false

• WB

Supplier Data

Western blot - Anti-TYK2 antibody [EPR24628-106] (AB303500)

Blocking and diluting buffer and concentration : 5% NFDM/TBST In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution. In Western blot, Anti-TYK2 antibody - Total protein control (ab303500) staining at 1/1000 dilution

All lanes:

Western blot - Anti-TYK2 (phospho Y1054 + Y1055) antibody [EPR28591-25] (<a href='/en-us/products/primary-antibodies/tyk2-phospho-y1054-y1055-antibody-epr28591-25-ab314878'>ab314878</a>) at 1/1000 dilution

Lane 1:

Untreated U266B1 (human multiple myeloma B lymphocyte) starved for 16 hours whole cell lysate (untreated membrane) at 20 µg

Lane 2:

U266B1 starved for 16 hours, then treated with 50ng/ml IFN alpha and 10μM MG-132 for 15 minutes whole cell lysate (untreated membrane) at 20 µg

Lane 3:

U266B1 starved for 16 hours, then treated with 50ng/ml IFN alpha and 10μM MG-132 for 15 minutes whole cell lysate (alkaline phosphatase treated membrane) 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

Observed band size: 135 kDa,37 kDa,140 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-TYK2 antibody [EPR24628-106] (AB303500)

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

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

All lanes:

Western blot - Anti-TYK2 antibody [EPR24628-106] (ab303500) at 1/1000 dilution

Lane 1:

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

Lane 2:

Neuro-2a (mouse neuroblastoma neuroblast), whole cell lysate at 20 µg

Lane 3:

U-87 MG (human glioblastoma-astrocytoma epithelial cell), whole cell lysate at 20 µg

Lane 4:

SH-SY5Y (human neuroblastoma 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 1/50000 dilution

Observed band size: 150 kDa

false

Exposure time: 81s

• WB

Lab

Western blot - Anti-TYK2 antibody [EPR24628-106] (AB303500)

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

All lanes:

Western blot - Anti-TYK2 antibody [EPR24628-106] (ab303500) at 1/1000 dilution

Lane 1:

Human liver tissue lysate at 40 µg

Lane 2:

Human kidney tissue lysate at 40 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 150 kDa

false

Exposure time: 114s

• WB

Lab

Western blot - Anti-TYK2 antibody [EPR24628-106] (AB303500)

Blocking and dilution buffer and concentration : Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS.

In Western blot, ab303500 was shown to bind specifically to TYK2. A band was observed at 150 kDa in wild-type 293T cell lysates with no signal observed at this size in TYK2 knockout cell line ab266730 (knockout cell lysate ab258743).

To generate this image, wild-type and TYK2 knockout 293T cell lysates were analyzed. First, samples were run on an SDS-PAGE gel then transferred onto an immobilon-FL PVDF membrane. Membranes were blocked in in Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS 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 (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

The samples were run on a Bis-Tris gel.

False colour image of Western blot : Anti-TYK2 antibody (ab303500) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red.

All lanes:

Western blot - Anti-TYK2 antibody [EPR24628-106] (ab303500) at 1/1000 dilution

Lane 1:

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

Lane 2:

TYK2 knockout 293T whole cell lysate at 20 µg

Lane 3:

Hela (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Secondary

Lanes 1 - 3:

Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed at 1/20000 dilution

Lanes 1 - 3:

Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed at 1/20000 dilution

Observed band size: 150 kDa

false

• WB

Lab

Western blot - Anti-TYK2 antibody [EPR24628-106] (AB303500)

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

All lanes:

Western blot - Anti-TYK2 antibody [EPR24628-106] (ab303500) at 1/1000 dilution

Lane 1:

Mouse lung tissue lysate at 40 µg

Lane 2:

Mouse kidney tissue lysate at 40 µg

Lane 3:

Rat lung tissue lysate at 40 µg

Lane 4:

Rat liver tissue lysate at 40 µg

Lane 5:

Rat kidney tissue lysate at 40 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 1/50000 dilution

Observed band size: 150 kDa

false

Exposure time: 114s