Anti-IL-2RG antibody [EPR24694-133]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-2RG antibody [EPR24694-133] (AB273023)

Immunohistochemical analysis of paraffin-embedded Human gastric cancer tissue labelling IL-2RG with ab273023 at 1/500 (1.126 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining in human gastric cancer. The section was incubated with ab273023 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-2RG antibody [EPR24694-133] (AB273023)

Immunohistochemical analysis of paraffin-embedded Human colon tissue labelling IL-2RG with ab273023 at 1/500 (1.126 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining in lamina propria of human colon. The section was incubated with ab273023 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• WB

Lab

Western blot - Anti-IL-2RG antibody [EPR24694-133] (AB273023)

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

The molecular weight observed is consistent with what has been described in the literature (PMID : 11418669 and PMID : 19329337). The 50kDa-70kDa bands correspond to the mature form and immature form.

Negative control : HT-29 (PMID : 8530527).

Exposure time : Lane 1 : 3 minutesLane 2 : 15 seconds Lanes 3-8 : 48 seconds

All lanes:

Western blot - Anti-IL-2RG antibody [EPR24694-133] (ab273023) at 1/1000 dilution

Lane 1:

Human spleen tissue lysate at 20 µg

Lane 2:

Human lymphoma tissue lysate at 20 µg

Lane 3:

Daudi (human Burkitts lymphoma lymphoblast), whole cell lysate at 20 µg

Lane 4:

Jurkat (human T cell leukemia T lymphocyte), whole cell lysate at 20 µg

Lane 5:

HT-29 (human colorectal adenocarcinoma epithelial cell), whole cell lysate at 20 µg

Lane 6:

RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage), whole cell lysate at 20 µg

Lane 7:

Mouse spleen tissue lysate at 20 µg

Lane 8:

Rat spleen tissue lysate at 20 µg

Secondary

Lanes 1 - 2:

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

Lanes 3 - 8:

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 1/20000 dilution

Predicted band size: 42 kDa

Observed band size: 50-70 kDa

false

• WB

Lab

Western blot - Anti-IL-2RG antibody [EPR24694-133] (AB273023)

Western blot : Anti-IL2RG antibody [EPR24694-133] (ab273023) 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, ab273023 was shown to bind specifically to IL2RG. A band was observed at 55 kDa in wild-type K562 cell lysates with no signal observed at this size in IL2RG knockout cell line. To generate this image, wild-type and IL2RG knockout K562 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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 IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-IL-2RG antibody [EPR24694-133] (ab273023) at 1/1000 dilution

Lane 1:

Wild-type K562 cell lysate at 20 µg

Lane 2:

IL2RG knockout K562 cell lysate at 20 µg

Lane 3:

Jurkat cell lysate at 20 µg

Lane 4:

MCF7 cell lysate at 20 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: 55 kDa

false

• IP

Supplier Data

Immunoprecipitation - Anti-IL-2RG antibody [EPR24694-133] (AB273023)

IL-2RG was immunoprecipitated from 0.35 mg Daudi (human Burkitt's lymphoma lymphoblast), whole cell lysate 10ug with ab273023 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab273023 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Daudi (human Burkitt's lymphoma lymphoblast), whole cell lysate 10ug

Lane 2 : ab273023 IP in Daudi whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab273023 in Daudi whole cell lysate

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

Exposure time : 75 seconds

All lanes:

Immunoprecipitation - Anti-IL-2RG antibody [EPR24694-133] (ab273023)

Predicted band size: 42 kDa

Observed band size: 50-70 kDa

false