Anti-Human IgM antibody [EPR20731]

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-Human IgM antibody [EPR20731] (AB212201)

Flow cytometric analysis of human PBMC (peripheral blood mononuclear cells) cell line labeling IgM with ab212201 at 1/50 (right panel) compared with an isotype control details (ab172730) (left panel). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077), at 1/2000 dilution was used as the secondary antibody.

Human PBMCs co-stained with anti-CD19 (Y-axis); only CD19+ population give a positive signal for IgM. Gated on total viable cells.

• WB

Lab

Western blot - Anti-Human IgM antibody [EPR20731] (AB212201)

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

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/1000000 dilution.

Low expression or negative samples : MOLT-4, Jurkat (PMID : 24141767, PMID : 26121639).

All lanes:

Western blot - Anti-Human IgM antibody [EPR20731] (ab212201) at 1/1000 dilution

Lane 1:

Raji (human Burkitt's lymphoma B lymphocyte) whole cell lysate at 20 µg

Lane 2:

Daudi (human Burkitt's lymphoma lymphoblast) whole cell lysate at 20 µg

Lane 3:

MOLT-4 (human lymphoblastic leukemia T lymphoblast) whole cell lysate at 20 µg

Lane 4:

Jurkat (human T cell leukemia T lymphocyte) 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 1/20000 dilution

Observed band size: 75 kDa

false

Exposure time: 5s

• WB

Supplier Data

Western blot - Anti-Human IgM antibody [EPR20731] (AB212201)

The blot was developed on a BIO-RAD® ChemiDoc™ MP instrument.
The molecular mass observed is consistent with the literature (PMID 16199094; PMID : 15955802; PMID 2107027).

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure times.

Lanes 1 & 3 : 3 minutes.

Lane 2 : 10 seconds.

All lanes:

Western blot - Anti-Human IgM antibody [EPR20731] (ab212201) at 1/1000 dilution

Lane 1:

Raji (human Burkitt's lymphoma cell line) whole cell lysate at 20 µg

Lane 2:

Human tonsil tissue lysate at 20 µg

Lane 3:

Human fetal spleen tissue lysate at 10 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/4000 dilution

Predicted band size: 49 kDa

Observed band size: 78 kDa

true

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Human IgM antibody [EPR20731] (AB212201)

Immunofluorescent analysis of 100% methanol-fixed Raji (human Burkitt's lymphoma cell line) cells labeling IgM with ab212201 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining in Raji cells.

The nuclear counterstain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) at 1/200 dilution (red).

Secondary antibody only control : Used PBS instead of primary antibody, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

• IP

Unknown

Immunoprecipitation - Anti-Human IgM antibody [EPR20731] (AB212201)

IgM was immunoprecipitated from 0.35 mg of Raji (human Burkitt's lymphoma cell line) whole cell lysate with ab212201 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab212201 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution.

Lane 1 : Raji whole cell lysate 10 μg (Input).

Lane 2 : ab212201 IP in Raji whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab212201 in Raji whole cell lysate.

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

Exposure time : 10 seconds.

All lanes:

Immunoprecipitation - Anti-Human IgM antibody [EPR20731] (ab212201)

Predicted band size: 49 kDa

Observed band size: 78 kDa

true