Description
The M18/2 antibody specifically recognizes the common β2 chain of LFA-1 (CD11a/CD18, αLβ2 integrin), Mac-1 (CD11b/CD18, αMβ2 integrin), and gp150, 95 (CD11c/CD18, αXβ2 integrin). Expression of CD18 is limited to leukocytes, where it is widely distributed in consort with the three integrin α chains (CD11a, CD11b, and CD11c). Among splenocytes, NK cells have the highest density of CD18, and T lymphocytes express a higher density than the remaining cells. The β2 integrins are important mediators of leukocyte-endothelium interactions. It has been reported that M18/2 antibody blocks in vivo metastasis of the LB lymphoma to the spleen and that it blocks in vitro formation of aggregates of LB cells and splenocytes. However, other reports indicate that mAb M18/2 has no effect on CTL-mediated killing, adherence of C3bi-sensitized erythrocytes to Mac-1, antigen-specific binding of T cells to antigen-producing cells, or rejection of cardiac allografts. Recent in vitro studies indicate that M18/2 antibody stimulates adhesion of Mac-1 to its ligands C3bi and ICAM-1, and it stimulates adhesion of LFA-1 to ICAM-1, but it has no effect upon the interactions of LFA-1 with ICAM-2 nor ICAM-3.
Bead-based compensation or unmixing controls, such as BD® CompBeads or BD™ SpectraComp™, can be used as surrogates to assess fluorescence spillover when bound to fluorochrome-conjugated antibodies. Although these beads have spectral properties similar to cells, variations in spectral emission may occur, resulting in differing spillover values compared to biological controls. Therefore, it is considered best practice to compare the spillover obtained from cells and bead-based controls when using BD® CompBeads or BD™ SpectraComp™ for the first time, to ensure they are appropriate for the intended application.
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to the dye under optimum conditions that minimize unconjugated dye and antibody.
1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
2. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
3. For U.S. patents that may apply, see bd.com/patents.
4. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
5. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
6. The production process underwent stringent testing and validation to assure that it generates a high-quality conjugate with consistent performance and specific binding activity. However, verification testing has not been performed on all conjugate lots.
7. When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
8. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
9. An isotype control should be used at the same concentration as the antibody of interest.
10. Please observe the following precautions: We recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to protect exposure of conjugated reagents, including cells stained with those reagents, to any room illumination. Absorption of visible light can significantly affect the emission spectra and quantum yield of tandem fluorochrome conjugates.
11. Alexa Fluor™ is a trademark of Life Technologies Corporation.
