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MEGACD40L® (soluble) (human), (recombinant)

High activity, high purity CD40L construct for co-stimulatory activation of an immune response
 
ALX-522-110-C010 10 µg 499.00 USD
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  • Highly active MEGACD40L® oligomer mimics in vivo membrane-assisted CD40L aggregation and stimulation without the need for enhancers
  • High purity with low endotoxin levels
  • Experienced manufacturing results in consistent and dependable lot-to-lot performance
  • Optimized formulation for improved stability and enhanced activity
MEGACD40L® is a high activity construct in which two trimeric CD40 ligand molecules are artificially linked via the collagen domain of Adiponectin/ACRP30/AdipoQ. This construct very effectively simulates the natural membrane-assisted aggregation of CD40L in vivo. It provides a simple and equally potent alternative to [CD40L+enhancer] combinations (Prod. No. ALX-850-064).

Product Specification

Alternative Name:CD40L:ACRP30headless, CD154:ACRP30headless, TNFSF 5:ACRP30headless, gp39:ACRP30headless
 
MW:~38kDa
 
Source:Produced in CHO cells. The extracellular domain of human CD40L (CD154) (aa 116-261) is fused at the N-terminus to mouse ACRP30headless (aa 18-111) and a FLAG®-tag.
 
UniProt ID:P29965 (human CD40L), Q60994 (mouse Adiponectin)
 
Concentration:0.1mg/ml after reconstitution.
 
Formulation:Lyophilized. Contains PBS.
 
Purity:≥90% (SDS-PAGE)
 
Purity Detail:Purified by multi-step chromatography.
 
Endotoxin Content:≤0.01EU/µg purified protein (LAL test; Cape Cod Associates).
 
Biological Activity:Binds to human CD40. Induces B cells activation (as demonstrated by dose-dependent upregulation of CD86).
 
Application Notes:Can be used to mediate immune and inflammatory responses, as a protein tool for pre-clinical studies in immunotherapy, to monitor patients undergoing chemotherapy and to mimic autoimmune phenotypes in vivo.
 
Reconstitution:Reconstitute with 100µl sterile water. Further dilutions should be made with medium containing 5% fetal calf serum.
 
Shipping:Shipped on Blue Ice
 
Long Term Storage:-20°C
 
Use/Stability:Stable for at least 6 months after receipt when stored at -20°C.
 
Handling:Avoid freeze/thaw cycles. After reconstitution, prepare aliquots and store at -20°C.
 
Technical Info/Product Notes:FLAG is a registered trademark of Sigma-Aldrich Co.
 
ALX-522-110 Fig1
Graphical representation of MEGACD40L®, Soluble (human) (recombinant) (Prod. No. ALX-522-110).
ALX-522-110 Fig6
PBMCs (peripheral blood mononuclear cells) were incubated at 37°C, 5% CO2 for 48 hours in 48 well plates (1 x 106 cells per well) containing 200µl serum free test media with serially diluted MEGACD40L® (Prod. No. ALX-522-110), CD40L (Prod. No. ALX-522-015), or CD40L + 2µg/ml Enhancer (Prod. No. ALX-804-034). After treatment the cells were washed 3X with PBS + 1% BSA. The cells were dual stained for 2 hours at 4°C with mouse anti-human CD19 (PE conjugate) and mouse anti-human CD86 (APC conjugate). The cells were washed 2X with PBS + 1% BSA, then re-suspended In PBS. Samples were analyzed on a BD Facs Calibur flow cytometer. The data is presented as the percent of CD86 positive cells per CD19 positive B cells at each concentration.
ALX-522-110 Fig5
MEGACD40L® was subjected to stability testing by storing the protein at the indicated temperature for 2 months. The protein was then tested for activity using a B cell activation assay. Results indicate MEGACD40L® retains activity under even harsh storage conditions.
ALX-522-110 Fig4
MEGACD40L® from 7 consecutive manufacturing lots was tested using a B-cell activation assay at 1000ng/mL.
ALX-522-110 Fig3
B cell lymphocyte activation by MEGACD40L® (Prod. No. ALX-522-110). Method: Peripheral Blood Mononuclear Cells (PBMCs) were incubated at 37°C, 5% CO2 for 48 hours in 48 well plates (1 x 106 cells/well). Each well contained 200µl serum free test media with serially diluted MEGACD40L®. After treatment, cells were washed and dual-stained with mouse anti-human CD19–PE and mouse anti-human CD86–APC and analyzed by flow cytometry. The data are presented as the percent of CD19 positive B cells that co-stain as CD86 positive, at each concentration of MEGACD40L®.
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ALX-522-110 Fig1 ALX-522-110 Fig6 ALX-522-110 Fig5 ALX-522-110 Fig4 ALX-522-110 Fig3

Product Literature References

Expression of HIV-1 matrix protein p17 and association with B-cell lymphoma in HIV-1 transgenic mice: V.A. Carroll, et al.; PNAS 113, 13168 (2016), Application(s): Human B cells culture, Abstract; Full Text
T-helper signals restore B-cell receptor signaling in autoreactive anergic B cells by upregulating CD45 phosphatase activity: P. Szodoray, et al.; J. Allergy Clin. Immunol. 138, 839 (2016), Application(s): Cell culture, Abstract;
A human monoclonal antibody targeting the conserved staphylococcal antigen IsaA protects mice against Staphylococcus aureus bacteremia: S. van den Berg, et al.; Int. J. Med. Microbiol. 305, 55 (2015), Application(s): ELISA, PCR of human blood samples, Abstract;
CD40L Induces Functional Tunneling Nanotube Networks Exclusively in Dendritic Cells Programmed by Mediators of Type 1 Immunity: C. Zaccard, et al.; J. Immunol. 194, 1047 (2015), Application(s): Activation of mature human dendritic cells, Abstract;
Phenotypic Approaches to Identify Inhibitors of B Cell Activation: E. B. Rex, et al.; J. Biomol. Screen. 20, 876 (2015), Application(s): Assay, Abstract; Full Text
A GPBAR1 (TGR5) small molecule agonist shows specific inhibitory effects on myeloid cell activation in vitro and reduces experimental autoimmune encephalitis (EAE) in vivo: N. Lewis, et al.; PLoS One 9, e100883 (2014), Application(s): Assay of human PBMC, Abstract; Full Text
Development of a novel anti-canine CD20 monoclonal antibody with diagnostic and therapeutic potential: D. Ito, et al.; Leuk. Lymphoma 56, 219 (2014), Application(s): Analysis of canine B-cell lymphoma cells, Abstract;
Exosomes Derived from Burkitt’s Lymphoma Cell Lines Induce Proliferation, Differentiation, and Class-Switch Recombination in B Cells: C. Gutzeit, et al.; J. Immunol. 192, 5852 (2014), Application(s): PCR, Assay of human B cells, Abstract; Full Text
Preclinical evaluation of the novel, orally bioavailable Selective Inhibitor of Nuclear Export (SINE) KPT-335 in spontaneous canine cancer: results of a phase I study: C. London, et al.; PLoS One 9, e87585 (2014), Application(s): Analysis of canine tumor cells by WB, FLOW, Abstract; Full Text
Thyrotropin receptor and CD40 mediate interleukin-8 expression in fibrocytes: implications for thyroid-associated ophthalmopathy (an American Ophthalmological Society thesis).: R. Douglas, et al.; Trans. Am. Ophthalmol. Soc. 12, 26 (2014), Application(s): In vitro analysis of human fibrocytes, Abstract; Full Text
AMPKα1 deficiency amplifies proinflammatory myeloid APC activity and CD40 signaling: K.C. Carroll, et al.; J. Leukoc. Biol. 94, 1113 (2013), Abstract;
Blood CD4+CD45RO+CXCR5+ T cells are decreased but partially functional in signal transducer and activator of transcription 3 deficiency: F. Mazerolles, et al.; J. Allergy Clin. Immunol. 131, 1146 (2013), Application(s): Assay of blood cells, Abstract;
Blood CD4+CD45RO+CXCR5+ T cells are decreased but partially functional in signal transducer and activator of transcription 3 deficiency: F. Mazerolles, et al.; J. Allergy Clin. Immunol. 131, 1146 (2013), Application(s): Assay of blood cells, Abstract;
CD40 ligand is necessary and suffi cient to support primary diff use large B-cell lymphoma cells in culture: a tool for in vitro preclinical studies with primary B-cell malignancies: D. Ito, et al.; Leuk. Lymphoma 53, 1390 (2012), Application(s): Flow cytometry of human B cells, Abstract; Full Text
CD40 ligand is necessary and sufficient to support primary diffuse large B-cell lymphoma cells in culture: a tool for in vitro preclinical studies with primary B-cell malignancies: D. Ito, et al.; Leuk. Lymphoma 53, 1390 (2012), Application(s): Cell Culture, Abstract;
HIV-1 gp120 impairs the induction of B cell responses by TLR9-activated plasmacytoid dendritic cells: N.P. Chung, et al.; J. Immunol. 189, 5257 (2012), Application(s): Activation of dendritic cells, Abstract; Full Text
Immunodeficiency, autoinflammation and amylopectinosis in humans with inherited HOIL-1 and LUBAC deficiency: B. Boisson, et al.; Nat. Immunol. 13, 1178 (2012), Application(s): ELISA of human blood cells, Abstract; Full Text
Interleukin-6 Production in CD40-Engaged Fibrocytes in Thyroid-Associated Ophthalmopathy: Involvement of Akt and NF-κB: E.F. Gillespie, et al.; Invest. Ophthalmol. Vis. Sci. 53, 7746 (2012), Application(s): CD40 ligation, induction and signaling in Graves' orbital fibroblasts (GOFB) and fibrocytes, Abstract; Full Text
Siglec-7 tetramers characterize B-cell subpopulations and leukemic blasts: F. Gieseke, et al.; Eur. J. Immunol. 42, 2176 (2012), Application(s): ELISA of human B cells , Abstract; Full Text
Effects of NOD-like receptors in human B lymphocytes and crosstalk between NOD1/NOD2 and Toll-like receptors: T. Petterson, et al.; J. Leukoc. Biol. 89, 177 (2011), Abstract;
New mechanism of X-linked anhidrotic ectodermal dysplasia with immunodeficiency: impairment of ubiquitin binding despite normal folding of NEMO protein: M. Hubeau, et al.; Blood 118, 926 (2011), Abstract;
Reconstitution of huPBL-NSG mice with donor-matched dendritic cells enables antigen-specific T-cell activation: A. Harui, et al.; J. Neuroimmune Pharmacol. 6, 148 (2011), Application(s): Activation of dendritic cells, Abstract; Full Text
Study of patients with Hyper-IgM type IV phenotype who recovered spontaneously during late childhood and review of the literature: N. Karaca, et al.; Eur. J. Pediatr. 170, 1039 (2011), Application(s): Determining human B-cell function in Hyper-IgM blood samples, Abstract;
The CD40-CD154 interaction would correlate with proliferation and immune escape in pancreatic ductal adenocarcinoma: Y. Shoji, et al.; J. Surg. Oncol. 106, 230 (2011), Abstract;
Human TRAF3 adaptor molecule deficiency leads to impaired Toll-like Receptor 3 response and susceptibility to Herpes Simplex Encephalitis: R. Perez de Diego, et al.; Immunity 33, 400 (2010), Abstract;
Reconstitution of huPBL-NSG mice with donor-matched dendritic cells enables antigen-specific T-cell activation: A. Harui, et al.; J. Neuroimmune Pharmacol. 6, 148 (2010), Abstract; Full Text
Therapeutic Implications of the IL-21: IL-4 receptor system in children with common variable immunodeficiency syndrome: S. Borte, et al.; Klin. Padiatr. 222, 362 (2010), Abstract;
Multimeric soluble CD40 ligand (sCD40L) efficiently enhances HIV specific cellular immune responses during DNA prime and boost with attenuated poxvirus ventors MVA and NYVAC expressing HIV antigens : C.E. Gomez, et al.; Vaccine 27, 3165 (2009), Abstract;
Small-molecule costimulatory blockage: organic dye inhibitors of the CD40-CD154 interaction: E. Margolles-Clark, et al.; J Mol Med 87, 1133 (2009), Abstract;
Impaired CD40L signaling is a cause of defective IL-12 and TNF-alpha production in Sezary syndrome: circumvention by hexameric soluble CD40L: L.E. French, et al.; Blood 105, 219 (2005), Abstract; Full Text
Two adjacent trimeric Fas ligands are required for Fas signaling and formation of a death-inducing signaling complex: N. Holler, et al.; Mol. Cell. Biol. 23, 1428 (2003), Abstract; Full Text

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