Anti-Bcr antibody [EPR22062] - BSA and Azide free

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Bcr antibody [EPR22062] - BSA and Azide free (AB237039)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized K562 (human chronic myelogenous leukemia cell line from bone marrow) cell line labeling Bcr with ab222406 at 1/500 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells incubated with secondary antibody only) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab222406).

• IP

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Immunoprecipitation - Anti-Bcr antibody [EPR22062] - BSA and Azide free (AB237039)

Bcr was immunoprecipitated from 0.35 mg HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab222406 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab222406 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1/1000 dilution.

Lane 1 : HeLa whole cell lysate 10μg (Input).
Lane 2 : ab222406 IP in HeLa whole cell lysate (+)
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab222406 in HeLa whole cell lysate (-).

Blocking and dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 10 seconds.

The 160 kDa and 130 kDa bands are isoforms of BCR (PMID : 2494632, PMID : 3078961).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab222406).

All lanes:

Immunoprecipitation - Anti-Bcr antibody [EPR22062] (<a href='/en-us/products/primary-antibodies/bcr-antibody-epr22062-ab222406'>ab222406</a>)

Predicted band size: 143 kDa

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• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Bcr antibody [EPR22062] - BSA and Azide free (AB237039)

Immunohistochemical analysis of paraffin-embedded rat cerebrum tissue labeling Bcr with ab222406 at 1/500 dilution, followed by Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use. Cytoplasmic staining in rat cerebrum (PMID : 25331951). Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use.

Heat-mediated antigen retrieval using ab208572 (Universal HIER antigen retrieval reagent).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab222406).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Bcr antibody [EPR22062] - BSA and Azide free (AB237039)

Immunohistochemical analysis of paraffin-embedded mouse hippocampus tissue labeling Bcr with ab222406 at 1/500 dilution, followed by Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use. Cytoplasmic staining in mouse hippocampus (PMID : 25331951). Counterstained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use.

Heat-mediated antigen retrieval using ab208572 (Universal HIER antigen retrieval reagent).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab222406).

• WB

Lab

Western blot - Anti-Bcr antibody [EPR22062] - BSA and Azide free (AB237039)

This data was developed using the same antibody clone in a different buffer formulation (ab222406).

Lanes 1 - 2 : Merged signal (red and green). Green - ab222406 observed at 150 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55kDa.

ab222406 was shown to react with BCR in wild-type HEK-293T cells in western blot. The bands observed in BCR CRISPR/Cas9 edited cell line ab266583 (BCR CRISPR/Cas9 edited cell lysate ab257858) below 150kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type and BCR CRISPR/Cas9 edited HEK-293T cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab222406 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4<sup>&

176;</sup>C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Bcr antibody [EPR22062] (<a href='/en-us/products/primary-antibodies/bcr-antibody-epr22062-ab222406'>ab222406</a>) at 1/1000 dilution

Lane 1:

Wild-type HEK-293T cell lysate at 20 µg

Lane 2:

BCR CRISPR/Cas9 edited HEK-293T cell lysate at 20 µg

Predicted band size: 143 kDa

Observed band size: 150 kDa

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