BD Pharmingen™ PE Mouse Anti-Mouse α-GalCer:CD1d Complex
Clone L363 (RUO)
Flow cytometric analysis of α-GalCer:CD1d complex expression on Mouse JAWSII cells. Cells from the Mouse JAWSII (Immature dendritic cell; ATCC® CRL-11904™) cell line were either not (Left Histograms) or were (Right Histograms) loaded overnight with α-GalCer. The cells were then preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142) and stained with either PE Mouse IgG2a, κ Isotype Control (Cat. No. 553457; dotted line histograms) or PE Mouse Anti-Mouse α-GalCer:CD1d complex antibody (Cat. No. 569977/569978; solid line histograms) at 0.5 μg/test. DAPI (4',6-Diamidino-2-Phenylindole, Dihydrochloride) Solution (Cat. No. 564907) was added to cells right before analysis. The fluorescence histograms showing α-GalCer:CD1d complex expression (or Ig Isotype control staining) were derived from gated events with the light-scatter characteristics of viable (DAPI-negative) cells. Flow cytometry and data analysis were performed using a BD LSRFortessa™ Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.
Flow cytometric analysis of α-GalCer:CD1d complex expression on Mouse JAWSII cells. Cells from the Mouse JAWSII (Immature dendritic cell; ATCC® CRL-11904™) cell line were either not (Left Histograms) or were (Right Histograms) loaded overnight with α-GalCer. The cells were then preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142) and stained with either PE Mouse IgG2a, κ Isotype Control (Cat. No. 553457; dotted line histograms) or PE Mouse Anti-Mouse α-GalCer:CD1d complex antibody (Cat. No. 569977/569978; solid line histograms) at 0.5 μg/test. DAPI (4',6-Diamidino-2-Phenylindole, Dihydrochloride) Solution (Cat. No. 564907) was added to cells right before analysis. The fluorescence histograms showing α-GalCer:CD1d complex expression (or Ig Isotype control staining) were derived from gated events with the light-scatter characteristics of viable (DAPI-negative) cells. Flow cytometry and data analysis were performed using a BD LSRFortessa™ Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.
Preparation And Storage
Recommended Assay Procedures
BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation). When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cell and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.
Product Notices
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- An isotype control should be used at the same concentration as the antibody of interest.
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Companion Products
The L363 monoclonal antibody specifically recognizes the complex formed by alpha-Galactosylceramide (α-Galactosylceramide) and Mouse CD1d, Mouse α-GalCer:CD1d Complex. The α-Galactosylceramide (α-GalCer) glycolipid can activate mouse and human iNKT cells. α-GalCer has been used as a model antigen for the analyses of iNKT cell-mediated immunity. This glycolipid binds to the nonclassical MHC I CD1d molecule expressed on antigen presenting cells such as dendritic cells. The cell-bound α-GalCer:CD1d Complex can subsequently be recognized by iNKT cells through their invariant Vα14-Jα18 TCR α chain. Activated iNKT cells have a broad array of effector and regulatory immune functions due to their rapid response to antigens by production of Th1 and Th2 cell-associated cytokines and activation of effector functions performed by other leucocytes. The L363 antibody can reportedly block activation of iNKT cells by the Mouse α-GalCer:CD1d Complex.
Development References (5)
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Arora P, Baena A, Yu KO, et al. A single subset of dendritic cells controls the cytokine bias of natural killer T cell responses to diverse glycolipid antigens.. Immunity. 2014; 40(1):105-16. (Clone-specific: Flow cytometry). View Reference
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Bai L, Sagiv Y, Liu Y, et al. Lysosomal recycling terminates CD1d-mediated presentation of short and polyunsaturated variants of the NKT cell lipid antigen alphaGalCer.. Proc Natl Acad Sci U S A. 2009; 106(25):10254-9. (Clone-specific: Flow cytometry, Fluorescence microscopy, Immunofluorescence). View Reference
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Im JS, Arora P, Bricard G, et al. Kinetics and cellular site of glycolipid loading control the outcome of natural killer T cell activation.. Immunity. 2009; 30(6):888-98. (Clone-specific: Flow cytometry, Fluorescence microscopy, Immunofluorescence). View Reference
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Yu ED, Girardi E, Wang J, et al. Structural basis for the recognition of C20:2-αGalCer by the invariant natural killer T cell receptor-like antibody L363.. J Biol Chem. 2012; 287(2):1269-78. (Clone-specific: ELISA). View Reference
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Yu KO, Im JS, Illarionov PA, et al. Production and characterization of monoclonal antibodies against complexes of the NKT cell ligand alpha-galactosylceramide bound to mouse CD1d.. J Immunol Methods. 2007; 323(1):11-23. (Immunogen: Flow cytometry). View Reference
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.
Refer to manufacturer's instructions for use and related User Manuals and Technical Data Sheets before using this product as described.
Comparisons, where applicable, are made against older BD technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.