Biolegend, 392902, Purified anti-human CD66b Antibody, 100μg

CD66b is a 95-100 kD glycosylphosphatidylinositol (GPI)-linked protein also known as CD67, CGM6, and NCA-95. CD66b is a member of the immunoglobulin superfamily, carcinoembryonic antigen (CEA)-like subfamily. CD66b, expressed on granulocytes, has been reported to induce activation in neutrophils and to be involved in heterophilic adhesion with CD66c.
100μg
Verified Reactivity: Human
Antibody Type: Monoclonal
Host Species: Mouse
Immunogen: Recombinant soluble human CEACAM8-Fc produced in HEK293 cells
Formulation: Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide.
Preparation: The antibody was purified by affinity chromatography.
Concentration: 0.5 mg/ml
Storage & Handling: The antibody solution should be stored undiluted between 2°C and 8°C.
Application: FC - Quality tested IHC-P - Verified SB - Community verified
Recommended Usage: Each lot of this antibody is quality control tested by immunofluorescent staining with flow cytometric analysis. For flow cytometric staining, the suggested use of this reagent is ≤ 0.5 µg per million cells in 100 µl volume. For immunohistochemistry, a concentration range of 5.0 - 10 µg/ml is suggested. It is recommended that the reagent be titrated for optimal performance for each application.
Additional Product Notes: For the use of this antibody in spatial biology applications, we have partnered with Lunaphore Technologies for demonstration of our antibodies on the COMET™. The COMET™ platform is an automated, end-to-end spatial biology solution developed for rapid and flexible multiplex tissue profiling. More information on the COMET™ and a complete list of our antibodies that have been demonstrated on the COMET™ can be found here.
Application References(PubMed link indicates BioLegend citation): Singer, et al. 2014. PLoS One. 9(4):e94106. PMID: 24743304 Klaile, et al. 2013. Respir Res. 14:85. PMID 23941132
Product Citations: Fox-Fisher I, et al. 2022. Med (N Y). 3:468. PubMed Vondra S, et al. 2023. Cell Rep. 42:111977. PubMed He M, et al. 2021. J Gynecol Oncol. e32:32. PubMed Riquelme E et al. 2019. Cell. 178(4):795-806 . PubMed Zheng Y, et al. 2022. iScience. 25:103785. PubMed
RRID: AB_2728422 (BioLegend Cat. No. 392902)
Structure: Ig superfamily, CEA antigen group, GPI-linked glycoprotein
Distribution: Granulocytes
Function: Cell adhesion, neutrophil activation
Ligand/Receptor: CD66c
Cell Type: Granulocytes
Biology Area: Immunology
Molecular Family: Adhesion Molecules, CD Molecules
Antigen References: Kuijpers T, et al. 1993. J Immunol. 151:4934. Kuroki M, et al. 1992. J Leuk Biol. 52:551.
Gene ID: 1088
UniProt: View information about CD66b on UniProt.org
Clone: 6/40c
Regulatory Status: RUO
Other Names: CEACAM8, CD67, CGM6, NCA-95
Isotype: Mouse IgG1, κ
Q: If an antibody clone has been previously successfully used in IBEX in one fluorescent format, will other antibody formats work as well?
A: It’s likely that other fluorophore conjugates to the same antibody clone will also be compatible with IBEX using the same sample fixation procedure. Ultimately a directly conjugated antibody’s utility in fluorescent imaging and IBEX may be specific to the sample and microscope being used in the experiment. Some antibody clone conjugates may perform better than others due to performance differences in non-specific binding, fluorophore brightness, and other biochemical properties unique to that conjugate.
Q: Will antibodies my lab is already using for fluorescent or chromogenic IHC work in IBEX?
A: Fundamentally, IBEX as a technique that works much in the same way as single antibody panels or single marker IF/IHC. If you’re already successfully using an antibody clone on a sample of interest, it is likely that clone will have utility in IBEX. It is expected some optimization and testing of different antibody fluorophore conjugates will be required to find a suitable format; however, legacy microscopy techniques like chromogenic IHC on fixed or frozen tissue is an excellent place to start looking for useful antibodies.
Q: Are other fluorophores compatible with IBEX?
A: Over 18 fluorescent formats have been screened for use in IBEX, however, it is likely that other fluorophores are able to be rapidly bleached in IBEX. If a fluorophore format is already suitable for your imaging platform it can be tested for compatibility in IBEX.
Q: The same antibody works in one tissue type but not another. What is happening?
A: Differences in tissue properties may impact both the ability of an antibody to bind its target specifically and impact the ability of a specific fluorophore conjugate to overcome the background fluorescent signal in a given tissue. Secondary stains, as well as testing multiple fluorescent conjugates of the same clone, may help to troubleshoot challenging targets or tissues. Using a reference control tissue may also give confidence in the specificity of your staining.
Q: How can I be sure the staining I’m seeing in my tissue is real?
A: In general, best practices for validating an antibody in traditional chromogenic or fluorescent IHC are applicable to IBEX. Please reference the Nature Methods review on antibody based multiplexed imaging for resources on validating antibodies for IBEX.