Expression of intracellular LIF by in-vitro-activated human peripheral blood cells. Human peripheral blood mononuclear cells (HPBMC) isolated by density gradient centrifugation (Ficoll-Paque™) were stimulated with plate-bound anti-human CD3 antibody (10 µg/ml, Cat. No. 555336) and soluble anti-CD28 antibody (2 µg/ml, Cat. No. 555725) in the presence of human IL-2 (10 ng/ml, Cat. No. 554603) and IL-4 (40 ng/ml, Cat. No. 554605) for 2 days. The cells were subsequently washed and expanded in IL-2 and IL-4 for 3 days. Following expansion, the cells were washed and stimulated for 5 hrs with PMA (5 ng/ml) and ionomycin (500 ng/ml) in the presence of BD GolgiPlug™ (Cat. No. 555029). Following incubation, the cells were harvested, washed and fixed with BD Cytofix/Cytoperm™ Solution (Cat. No. 554722, 15 min, 4°C). The intracellular levels of LIF expressed by activated HPBMC were detected by immunofluorescent staining and flow cytometric analysis using the PE-conjugated 1F10 antibody (left panel, 20 µl/10e6 cells, Cat. No. 558571) or an immunoglobulin isotype control (right panel). Dot plots (left and right panels) were derived from gated events with the forward- and side- light scatter characteristics of mononuclear cells.
Expression of intracellular LIF by in-vitro-activated human peripheral blood cells. Human peripheral blood mononuclear cells (HPBMC) isolated by density gradient centrifugation (Ficoll-Paque™) were stimulated with plate-bound anti-human CD3 antibody (10 µg/ml, Cat. No. 555336) and soluble anti-CD28 antibody (2 µg/ml, Cat. No. 555725) in the presence of human IL-2 (10 ng/ml, Cat. No. 554603) and IL-4 (40 ng/ml, Cat. No. 554605) for 2 days. The cells were subsequently washed and expanded in IL-2 and IL-4 for 3 days. Following expansion, the cells were washed and stimulated for 5 hrs with PMA (5 ng/ml) and ionomycin (500 ng/ml) in the presence of BD GolgiPlug™ (Cat. No. 555029). Following incubation, the cells were harvested, washed and fixed with BD Cytofix/Cytoperm™ Solution (Cat. No. 554722, 15 min, 4°C). The intracellular levels of LIF expressed by activated HPBMC were detected by immunofluorescent staining and flow cytometric analysis using the PE-conjugated 1F10 antibody (left panel, 20 µl/10e6 cells, Cat. No. 558571) or an immunoglobulin isotype control (right panel). Dot plots (left and right panels) were derived from gated events with the forward- and side- light scatter characteristics of mononuclear cells.
Product Details
BD Pharmingen™
MLPLI; HILDA; HSF III
Human (QC Testing)
Mouse IgG1, κ
Recombinant vaccinia virus
Intracellular staining (flow cytometry) (Routinely Tested)
20 µl
AB_397085
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO
Preparation And Storage
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 with R-PE under optimum conditions, and unconjugated antibody and free PE were removed.
Product Notices
- This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Ficoll-Paque is a trademark of Amersham Biosciences Limited.
- 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.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- 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.
558571 Rev. 3
Antibody Details
1F10
The 1F10 antibody specifically binds to human Leukemia Inhibitory Factor (LIF) also known as Human Interleukin for DA cells (HILDA), Melanoma-derived Lipoprotein Lipase Inhibitor (MLPLI) and Hepatocyte Stimulating Factor III (HSF III). LIF is produced by multiple sources including activated T cells and macrophages, myelomonocytic lineages, fibroblasts, liver, heart and melanoma cells. LIF regulates the differentiation of embryonic stem cells, neural cells, osteoblasts, adipocytes, hepatocytes and kidney epithelial cells. Other activities include terminal differentiation in leukemic cells and the stimulation of acute-phase protein synthesis in hepatocytes. Many of its biological functions parallel those of Interleukin-6, Oncostatin M, Ciliary Neurotrophic Factor, Interleukin-11 and Cardiotrophin-1. In vivo LIF is important in regulating the inflammatory response by tuning the balance of four systems in the body, namely the immune, the haematopoietic, the nervous and the endocrine systems. The immunogen used to generate the 1F10 hybridoma was recombinant vaccinia virus encoding the human LIF cytokine. The LIF cytokine had been engineered to get expressed on the membrane of the infected cell. The attachment to the membrane was obtained with the glycosylphoshosphatidyl anchor targeting DNA sequence from the DAF molecule (CD55).
558571 Rev. 3
Format Details
PE
R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
PE
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
576 nm
558571 Rev.3
Citations & References
Development References (12)
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Baumann H, Won KA, Jahreis GP. Human hepatocyte-stimulating factor-III and interleukin-6 are structurally and immunologically distinct but regulate the production of the same acute phase plasma proteins. J Biol Chem. 1989; 264(14):8046-8051. (Biology). View Reference
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Baumann H, Wong GG. Hepatocyte-stimulating factor III shares structural and functional identity with leukemia-inhibitory factor. J Immunol. 1989; 143(4):1163-1167. (Biology). View Reference
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Escary JL, Perreau J, Dumenil D, Ezine S, Brulet P. Leukaemia inhibitory factor is necessary for maintenance of haematopoietic stem cells and thymocyte stimulation. Nature. 1993; 363(6427):361-364. (Biology). View Reference
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Gearing DP. Leukemia inhibitory factor: does the cap fit. Ann N Y Acad Sci. 1991; 628:9-18. (Biology). View Reference
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Gough NM, Gearing DP, King JA, et al. Molecular cloning and expression of the human homologue of the murine gene encoding myeloid leukemia-inhibitory factor. Proc Natl Acad Sci U S A. 1988; 85(8):2623-2627. (Biology). View Reference
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Moreau JF, Donaldson DD, Bennett F, Witek-Giannotti J, Clark SC, Wong GG. Leukaemia inhibitory factor is identical to the myeloid growth factor human interleukin for DA cells. Nature. 1988; 336(6200):690-692. (Biology). View Reference
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Mori M, Yamaguchi K, Abe K. Purification of a lipoprotein lipase-inhibiting protein produced by a melanoma cell line associated with cancer cachexia. Biochem Biophys Res Commun. 1989; 160(3):1085-1092. (Biology). View Reference
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Taupin JL, Acres B, Dott K, et al. Immunogenicity of HILDA/LIF either in a soluble or in a membrane anchored form expressed in vivo by recombinant vaccinia viruses. Scand J Immunol. 1993; 38(3):293-301. (Biology). View Reference
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Taupin JL, Gualde N, Moreau JF. A monoclonal antibody based elisa for quantitation of human leukaemia inhibitory factor. Cytokine. 1997; 9(2):112-118. (Biology). View Reference
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Taupin JL, Pitard V, Dechanet J, Miossec V, Gualde N, Moreau JF. Leukemia inhibitory factor: part of a large ingathering family. Int Rev Immunol. 1989; 16(3-4):397-426. (Biology). View Reference
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Williams RL, Hilton DJ, Pease S, et al. Myeloid leukaemia inhibitory factor maintains the developmental potential of embryonic stem cells. Nature. 1988; 336(6200):684-687. (Biology). View Reference
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Yamamori T, Fukada K, Aebersold R, Korsching S, Fann MJ, Patterson PH. The cholinergic neuronal differentiation factor from heart cells is identical to leukemia inhibitory factor. Science. 1989; 246(4936):1412-1416. (Biology). View Reference
558571 Rev. 3
Please refer to Support Documents for Quality Certificates
Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.
