CD11b Monoclonal Antibody (M1/70), NovaFluor™ Blue 760, eBioscience™

Product Specific Information

Description
The M1/70 monoclonal antibody reacts with mouse CD11b, the 165-170 kDa integrin alphaM. CD11b non-covalently associates with CD18 to form alphaMbeta2 integrin (Mac-1) and binds to CD54 (ICAM-1), C3bi, and fibrinogen. Mac-1 is expressed by macrophages, NK cells, granulocytes, activated lymphocytes and mouse B-1 cells in the peritoneal cavity. M1/70 is also cross-reactive to human CD11b, and can be used for the detection of this antigen on human peripheral blood monocytes, granulocytes, and a subset of NK cells. Through interactions with its ligands, CD11b participates in adhesive cell interactions.

This product contains 1 vial of NovaFluor conjugate and 1 vial of CellBlox Plus Blocking Buffer.

Applications Tested
The M1/70 antibody has been tested by flow cytometric analysis of mouse splenocytes and bone marrow cells. This can be used at less than or equal to 0.4 µg per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. It is recommended that the antibody be carefully titrated for optimal performance in the assay of interest.

Master Mixes
• Master mixes of NFs should be made at 2-8 °C and may be made up to 4 hours ahead of time.
• We do not recommend storing master mixes containing NovaFluor conjugates overnight or longer.

Whole Blood Compatibility
• When utilizing whole blood (as opposed to density-gradient-purified PBMC), we recommend lysing red blood cells in bulk prior to staining with NovaFluor conjugates.
• See the Bulk Lysis of Human Whole Blood protocol here.
• Staining of whole blood with NovaFluor conjugates followed by lysis of red blood cells may result in higher-than-expected background staining.

Viability Dye Compatibility
• NovaFluor dyes are not compatible with DNA intercalcating dyes such as propidium iodide, 7-actinomycin D (7-AAD) and DAPI. Invitrogen LIVE/DEAD Fixable Dead Cell stains are recommended for use with NovaFluor dyes.

CellBlox Plus Blocking Buffer
• This NovaFluor conjugate comes with CellBlox Plus Blocking Buffer (Cat. No. C001T03F01), essential for optimal staining.
• Use CellBlox Plus Blocking Buffer in all experiments with NovaFluor conjugates.
• Add 5 μL per sample to antibody cocktails/master mixes (regardless of how many Novafluor-conjugated antibodies are present) before combining with cells.
• CellBlox Plus Blocking Buffer is compatible with either Super Bright Complete Blocking Buffer or Brilliant Stain Buffer and can be used in antibody cocktails/master mixes with those reagents.
• For single-color controls, use 5 μL of CellBlox Plus Blocking Buffer per 100 μL of cell sample (10^3 to 10^8 cells).
• CellBlox Plus Blocking Buffer may also be used with non-NovaFluor dyes to reduce non-specific binding of monocytes to tandem cyanine-based dyes.

NovaFluor conjugates are based on Phiton technology utilizing novel fluorophore-containing nucleic acid dye structures that allow for engineered fluorescent signatures with consideration for spillover and spread impacts. Learn more

Excitation: 492 nm; Emission: 766 nm; Laser: 488 nm (Blue) Laser

Target Information

CD11b (integrin alpha-M, ITGAM, integrin alpha-X, ITGAX) is a 165 kDa adhesion molecule that associates non-covalently with integrin beta-2 (CD18). The CD11b/CD18 heterodimeric complex is also known as integrin alpha-M beta-2, Mac-1, and CR3 (complement receptor 3). CD11b is expressed on the surface of monocytes/macrophages, granulocytes, activated lymphocytes, a subset of NK cells, a subset of dendritic cells, and microglia in the brain. CD11b/CD18 functions as the receptor for ICAM-1 (CD54), ICAM-2 (CD102), ICAM-4 (CD242), CD14, CD50, CD23, heparin, iC3b, fibrinogen, and Factor X -these adhesions are critical for cell-cell and cell-matrix interactions. CD11b is expressed on 8% of spleen cells, 44% of bone marrow cells, and less than 1% of thymocytes, and is commonly used as a microglial marker in nervous tissue. The expression of CD11b increases during monocyte maturation and expression levels vary on tissue macrophages. Further, peritoneal macrophages are reported to express higher levels of CD11b than splenic macrophages. Diseases associated with CD11b dysfunction include systemic lupus erythematosus 6 and ITGAM-related susceptibility to systemic lupus erythematosus.