Anti-IL-2RG antibody [EPR24694-133] - BSA and Azide free

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IL-2RG antibody [EPR24694-133] - BSA and Azide free (AB282585)

This data was developed using ab273023, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Human gastric cancer tissue labeling IL-2RG with ab273023 at 1/500 (1.126 ug/ml) followed by a Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) at Ready to use dilution. 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 Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) at Ready to use dilution.

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] - BSA and Azide free (AB282585)

This data was developed using ab273023, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Human colon tissue labeling IL-2RG with ab273023 at 1/500 (1.126 ug/ml) followed by a Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) at Ready to use dilution. 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 Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) at Ready to use dilution.

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

• IP

Supplier Data

Immunoprecipitation - Anti-IL-2RG antibody [EPR24694-133] - BSA and Azide free (AB282585)

This data was developed using ab273023, the same antibody clone in a different buffer formulation.

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] (<a href='/en-us/products/primary-antibodies/il-2rg-antibody-epr24694-133-ab273023'>ab273023</a>)

Predicted band size: 42 kDa

Observed band size: 50-70 kDa

false

• WB

Lab

Western blot - Anti-IL-2RG antibody [EPR24694-133] - BSA and Azide free (AB282585)

This data was developed using 273023, the same antibody clone in a different buffer formulation.

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] (<a href='/en-us/products/primary-antibodies/il-2rg-antibody-epr24694-133-ab273023'>ab273023</a>) 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] - BSA and Azide free (AB282585)

This data was developed using the same antibody clone in a different buffer formulation (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] (<a href='/en-us/products/primary-antibodies/il-2rg-antibody-epr24694-133-ab273023'>ab273023</a>) 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