Anti-AICDA antibody [5G9] - BSA and Azide free

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-AICDA antibody [5G9] - BSA and Azide free (AB255769)

Intracellular flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized K-562 (human chronic myelogenous leukemia lymphoblast, Left) / Ramos (human Burkitt's lymphoma B lymphocyte, Right) cells labelling AICDA with ab252813 at 0.316μg/mL (Red) compared with a rat monoclonal IgG (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rat IgG (Alexa Fluor^® 488, ab150157) at 1/2000 dilution was used as the secondary antibody. Negative control : K-562.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab252813).

• WB

Lab

Western blot - Anti-AICDA antibody [5G9] - BSA and Azide free (AB255769)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab252813). False colour image of Western blot : Anti-AICDA antibody [5G9] staining at 1.264 ug/ml, shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (ab181602) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab252813 was shown to bind specifically to AICDA. A band was observed at 22 kDa in wild-type Raji cell lysates with no signal observed at this size in AICDA knockout cell line ab277185 (knockout cell lysate ab277227). To generate this image, wild-type and AICDA knockout Raji 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-Rat IgG H&L (IRDye® 800CW) preabsorbed (ab253031) and Goat anti-Rabbit IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-AICDA antibody [5G9] (<a href='/en-us/products/primary-antibodies/aicda-antibody-5g9-ab252813'>ab252813</a>) at 1.264 µg/mL

Lane 1:

Wild-type Raji cell lysate at 20 µg

Lane 2:

AICDA knockout Raji cell lysate at 20 µg

Lane 3:

Daudi cell lysate at 20 µg

Lane 4:

SH-SY5Y cell lysate at 20 µg

Predicted band size: 23 kDa

Observed band size: 22 kDa

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• WB

Lab

Western blot - Anti-AICDA antibody [5G9] - BSA and Azide free (AB255769)

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

Negative control : K562 (PMID : 27217538).

Exposure time : 59 seconds.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab252813).

All lanes:

Western blot - Anti-AICDA antibody [5G9] (<a href='/en-us/products/primary-antibodies/aicda-antibody-5g9-ab252813'>ab252813</a>) at 1.264 µg/mL

Lane 1:

NAMALWA (human Burkitts lymphoma B lymphocyte), whole cell lysate at 10 µg

Lane 2:

Raji (human Burkitts lymphoma B lymphocyte), whole cell lysate at 10 µg

Lane 3:

K562 (human chronic myelogenous leukemia lymphoblast), whole cell lysate at 10 µg

Lane 4:

WEHI-231 (mouse B cell lymphoma B lymphocyte), whole cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rat IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rat-igg-h-l-hrp-ab205720'>ab205720</a>) at 1/2000 dilution

Predicted band size: 23 kDa

Observed band size: 23 kDa

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