Overview

  • Product name
    Human CD40L ELISA Kit (TNFSF5)
    See all CD40L kits
  • Detection method
    Colorimetric
  • Precision
    Intra-assay
    Sample n Mean SD CV%
    supernatant 5 3.8%
    Inter-assay
    Sample n Mean SD CV%
    supernatant 3 2.8%
  • Sample type
    Cell culture supernatant, Serum, Plasma
  • Assay type
    Sandwich (quantitative)
  • Sensitivity
    2.9 pg/ml
  • Range
    12.5 pg/ml - 800 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture supernatant 95 84% - 105%
    Serum 96 93% - 100%
    Heparin Plasma 106 102% - 112%
    EDTA Plasma 96 94% - 99%
    Citrate Plasma 104 96% - 119%

  • Assay time
    1h 30m
  • Assay duration
    One step assay
  • Species reactivity
    Reacts with: Human
    Does not react with: Mouse
  • Product overview

    Human CD40L SimpleStep ELISA® kit (ab196268) has been redeveloped with new capture and detector antibodies. While this does not affect the specificity of the results, it may result in increased sensitivity and will improve the consistency in the performance of the kit. 


    CD40L in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of CD40L protein in human serum, plasma, and cell culture supernatant samples


    The SimpleStep ELISA® employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. TMB Development Solution is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.


    CD40L (CD40 Ligand) is a glycoprotein belonging to the TNF family. CD40 Ligand is expressed predominantly on activated CD4 T lymphocytes and is also found in other cells including NK cells, mast cells, basophils, and eosinophils. It is involved in B cell proliferation and IgG production. CD40 Ligand is also involved in immunoglobulin class switching.


    Sensitivity in Sample Diluent 25BS: 6.2 pg/mL. Sensitivity in Sample Diluent NS: 2.9 pg/mL. 

  • Tested applications
    Suitable for: Sandwich ELISAmore details
  • Platform
    Microplate

Properties

  • Storage instructions
    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests
    10X Wash Buffer PT (ab206977) 1 x 20ml
    Antibody Diluent 4BI 1 x 6ml
    10X Human CD40L Capture Antibody 1 x 600µl
    10X Human CD40L Detector Antibody 1 x 600µl
    Human CD40L Lyophilized Recombinant Protein 2 vials
    Plate Seals 1 unit
    Sample Diluent 25BS 1 x 20ml
    Sample Diluent NS (ab193972) 1 x 50ml
    SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit
    Stop Solution 1 x 12ml
    TMB Development Solution 1 x 12ml
  • Research areas
  • Function
    Mediates B-cell proliferation in the absence of co-stimulus as well as IgE production in the presence of IL-4. Involved in immunoglobulin class switching.
    Release of soluble CD40L from platelets is partially regulated by GP IIb/IIIa, actin polymerization, and an matrix metalloproteinases (MMP) inhibitor-sensitive pathway.
  • Tissue specificity
    Specifically expressed on activated CD4+ T-lymphocytes.
  • Involvement in disease
    Defects in CD40LG are the cause of X-linked immunodeficiency with hyper-IgM type 1 (HIGM1) [MIM:308230]; also known as X-linked hyper IgM syndrome (XHIM). HIGM1 is an immunoglobulin isotype switch defect characterized by elevated concentrations of serum IgM and decreased amounts of all other isotypes. Affected males present at an early age (usually within the first year of life) recurrent bacterial and opportunistic infections, including Pneumocystis carinii pneumonia and intractable diarrhea due to cryptosporidium infection. Despite substitution treatment with intravenous immunoglobulin, the overall prognosis is rather poor, with a death rate of about 10% before adolescence.
  • Sequence similarities
    Belongs to the tumor necrosis factor family.
  • Post-translational
    modifications
    The soluble form derives from the membrane form by proteolytic processing.
    N-linked glycan is a mixture of high mannose and complex type. Glycan structure does not influence binding affinity to CD40.
    Not O-glycosylated.
  • Cellular localization
    Secreted and Cell membrane.
  • Information by UniProt
  • Alternative names
    • CD154
    • CD40 antigen ligand
    • CD40 ligand
    • CD40-L
    • CD40L
    • CD40L_HUMAN
    • CD40LG
    • gp39
    • hCD40L
    • HIGM1
    • IGM
    • IMD3
    • soluble form
    • T B cell activating molecule
    • T BAM
    • T cell antigen Gp39
    • T-cell antigen Gp39
    • TNF related activation protein
    • TNF-related activation protein
    • TNFSF5
    • TrAP
    • Tumor necrosis factor (ligand) superfamily member 5
    • Tumor necrosis factor (ligand) superfamily, member 5 (hyper-IgM syndrome)
    • Tumor necrosis factor ligand superfamily member 5
    see all
  • Database links

Applications

Our Abpromise guarantee covers the use of ab196268 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Sandwich ELISA Use at an assay dependent concentration.

Images

  • ELISA Protocol Summary
  • Standard Curve comparison between human CD40L SimpleStep ELISA kit and traditional ELISA kit from leading competitor. SimpleStep ELISA kit shows increased sensitivity.

  • Standard Curve comparison between current human CD40L SimpleStep ELISA kit and original ELISA kit. The current SimpleStep ELISA kit shows increased sensitivity.

  • The CD40L standard curve was prepared as described in Section 10. Raw data values are shown in the table. Background-subtracted data values (mean +/- SD) are graphed.

  • The CD40L standard curve was prepared as described in Section 10. Raw data values are shown in the table. Background-subtracted data values (mean +/- SD) are graphed.

  • Unstimulated PBMC cell culture supernatant samples were also measured; all values were below the detectable range of the assay. The concentrations of CD40L were measured in duplicates, interpolated from the CD40L standard curves and corrected for sample dilution. Undiluted samples are as follows: stimulated PBMC supernatant 100%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean CD40L concentration was determined to be 488 pg/ml in stimulated PBMC supernatant.

  • Interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean CD40L concentration in healthy serum was determined to be 34 pg/mL with a range of ND (non-detectable) – 37.6 pg/mL. The mean CD40L concentration in rheumatoid arthritis serum was determined to be 607 pg/mL.

     

     

Protocols

References

ab196268 has not yet been referenced specifically in any publications.

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