Anti-CD11b antibody [EPR19387]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-CD11b antibody [EPR19387] (AB184308)

Immunofluorescent analysis of 100% methanol-fixed RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) and WEHI-231 (Mouse B cell lymphoma cell line) cells labeling CD11b with ab184308 at 1/500 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1,000 dilution (green).

Confocal image showing membrane staining on RAW 264.7 cells and no staining on WEHI-231 cells (negative cell line).

The nuclear counter stain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1,000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) preadsorbed (ab150120) at 1/1,000 dilution (red).

The negative controls are as follows :
-ve control 1 : ab184308 at 1/500 dilution followed by ab150120 at 1/1,000 dilution.
-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1,000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-CD11b antibody [EPR19387] (AB184308)

CD11b was immunoprecipitated from 0.35 mg of RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate with ab184308 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab184308 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : RAW 264.7 whole cell lysate, 10μg (Input).

Lane 2 : ab184308 IP in RAW 264.7 whole cell lysate.

Lane 3 : Rabbit IgG,monoclonal [EPR25A]- Isotype Control (ab172730) instead of ab184308 in RAW 264.7 whole cell lysate.

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

Exposure time : 8 seconds.

All lanes:

Immunoprecipitation - Anti-CD11b antibody [EPR19387] (ab184308)

Predicted band size: 127 kDa

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• WB

Supplier Data

Western blot - Anti-CD11b antibody [EPR19387] (AB184308)

Blocking/Dilution buffer : 5% NFDM/TBST.

CD11b is a 170 kD glycoprotein also known as αM integrin, Mac-1 α subunit (PMID : 10603370). It is exclusively restricted to cells of the myeloid lineages and monocytes, but not in T or B cell lines, so WEHI-231 cell lysate serves as a negative control (PMID : 2457584).

All lanes:

Western blot - Anti-CD11b antibody [EPR19387] (ab184308) at 1/1000 dilution

Lane 1:

J774A.1 (Mouse macrophage reticulum cell sarcoma cell line) whole cell lysate at 20 µg

Lane 2:

M1 (Mouse myeloblast myeloid leukemia cell line) whole cell lysate at 20 µg

Lane 3:

WEHI-231 (Mouse B cell lymphoma cell line) whole cell lysate at 20 µg

Lane 4:

RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) 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 1/100000 dilution

Predicted band size: 127 kDa

Observed band size: 170 kDa

false

Exposure time: 3min