Human CD40 Ligand/TNFSF5 Quantikine ELISA Kit

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Human CD40 Ligand/TNFSF5 ELISA Standard Curve
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Human CD40 Ligand/TNFSF5 Quantikine ELISA Kit Summary

Assay Type
Solid Phase Sandwich ELISA
96-well strip plate
Assay Length
4.5 hours
Sample Type & Volume Required Per Well
Cell Culture Supernates (100 uL), Serum (20 uL), Platelet-poor EDTA Plasma (100 uL), Platelet-poor Heparin Plasma (100 uL)
10.1 pg/mL
Assay Range
62.5 - 4,000 pg/mL (Cell Culture Supernates, Serum, EDTA Plasma, Platelet-poor EDTA Plasma, Platelet-poor Heparin Plasma)
Natural and recombinant human CD40 Ligand
< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.
No significant interference observed with available related molecules.

Product Summary

The Quantikine Human CD40 Ligand Immunoassay is a 4.5 hour solid-phase ELISA designed to measure human CD40 Ligand in cell culture supernates, serum, and plasma. It contains E. coli-expressed recombinant human CD40 Ligand and has been shown to accurately quantitate the recombinant factor. Results obtained using natural human CD40 Ligand showed linear curves that were parallel to the standard curves obtained using the Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for naturally occurring human CD40 Ligand.


Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested on one plate to assess intra-assay precision
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in separate assays to assess inter-assay precision

Cell Culture Supernates, Serum, Platelet-poor EDTA Plasma, Platelet-poor Heparin Plasma

Intra-Assay Precision Inter-Assay Precision
Sample 1 2 3 1 2 3
n 20 20 20 40 40 40
Mean (pg/mL) 430 1212 2638 437 1205 2612
Standard Deviation 21.8 54.4 143 27.9 72.7 162
CV% 5.1 4.5 5.4 6.4 6 6.2


The recovery of CD40 Ligand spiked to levels throughout the range of the assay in various matrices was evaluated.

Sample Type Average % Recovery Range %
Cell Culture Media (n=4) 102 90-114
Platelet-poor EDTA Plasma (n=5) 98 86-107
Platelet-poor Heparin Plasma (n=5) 94 86-113


To assess the linearity of the assay, samples containing and/or spiked with high concentrations of CD40 Ligand were serially diluted with Calibrator Diluent to produce samples with values within the dynamic range of the assay.
Human CD40 Ligand/TNFSF5 ELISA Linearity

Scientific Data

Human CD40 Ligand/TNFSF5 ELISA Standard Curve

Product Datasheets

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Preparation and Storage

The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: CD40 Ligand/TNFSF5

CD40 Ligand, also known as TNFSF5, CD154, TRAP, and gp39, is a trimeric protein that is expressed in transmembrane and soluble forms. It is expressed by many immune cell types as well as by endothelial and smooth muscle cells of the vasculature. It binds to CD40 on antigen presenting cells, platelets, and endothelial and epithelial cells, leading to enhanced B cell activation and T cell dependent humoral immune responses. CD40 Ligand dysregulation contributes to immune pathology in AIDS, atherosclerosis, atherothrombosis, and restenosis.

Entrez Gene IDs:
959 (Human); 21947 (Mouse); 84349 (Rat)
Alternate Names:
CD154 antigen; CD154; CD40 antigen ligand; CD40 Ligand; CD40L; CD40-L; CD40LG; CD40LIGM; gp39; hCD40L; HIGM1; T-B cell-activating molecule; T-BAM; T-cell antigen Gp39; TNF-related activation protein; TNFSF5; TNFSF5IMD3; TRAP; TRAPtumor necrosis factor (ligand) superfamily, member 5 (hyper-IgM syndrome); tumor necrosis factor (ligand) superfamily member 5; Tumor necrosis factor ligand superfamily member 5
⚠ WARNING: This product can expose you to chemicals including N,N-Dimethylforamide, which is known to the State of California to cause cancer. For more information, go to

Assay Procedure

Refer to the product for complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.
  1.   Prepare all reagents, standard dilutions, and samples as directed in the product insert.
  2.   Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

  3. 100 µL Assay Diluent
  4.   Add 100 µL of Assay Diluent to each well.

  5. 100 µL Standard, Control, or Sample
  6.   Add 100 µL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours on a horizontal orbital microplate shaker.
  7.   Aspirate each well and wash, repeating the process 3 times for a total of 4 washes.

  8. 200 µL Conjugate
  9.   Add 200 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours on the shaker.
  10.   Aspirate and wash 4 times.

  11. 200 µL Substrate Solution
  12.   Add 200 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes on the benchtop. PROTECT FROM LIGHT.

  13. 50 µL Stop Solution
  14.   Add 50 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.

Citations for Human CD40 Ligand/TNFSF5 Quantikine ELISA Kit

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

42 Citations: Showing 1 - 10
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  1. B cell and monocyte phenotyping: A quick asset to investigate the immune status in patients with IgA nephropathy
    Authors: S Sendic, L Mansouri, S Lundberg, A Nopp, SH Jacobson, J Lundahl
    PLoS ONE, 2021;16(3):e0248056.
    Species: Human
    Sample Types: Plasma
  2. Disturbed Lipid Metabolism in Diabetic Patients with Manifest Coronary Artery Disease Is Associated with Enhanced Inflammation
    Authors: K Buschmann, Y Gramlich, R Chaban, M Oelze, U Hink, T Münzel, H Treede, A Daiber, GD Duerr
    International Journal of Environmental Research and Public Health, 2021;18(20):.
    Species: Human
    Sample Types: Serum
  3. Analysis of Genetic Variation in CD40 and CD40L: Relationship with mRNA Relative Expression and Soluble Proteins in Acute Coronary Syndrome
    Authors: DE Martínez-F, JR Padilla-Gu, F Casillas-M, E Valdés-Alv, B Parra-Reyn, M Aceves-Ram, JF Muñoz-Vall, U Zalapa Flo, JC Chávez Her, Y Valle
    J Immunol Res, 2019;2019(0):8063983.
    Species: Human
    Sample Types: Plasma
  4. Biological variation of immunological blood biomarkers in healthy individuals and quality goals for biomarker tests
    Authors: N Aziz, R Detels, JJ Quint, D Gjertson, T Ryner, AW Butch
    BMC Immunol., 2019;20(1):33.
    Species: Human
    Sample Types: Serum
  5. Multilevel defects in the hematopoietic niche in essential thrombocythemia
    Authors: T Sun, M Ju, X Dai, H Dong, W Gu, Y Gao, R Fu, X Liu, Y Huang, W Liu, Y Chi, W Wang, H Li, Y Zhou, L Shi, R Yang, L Zhang
    Haematologica, 2019;0(0):.
    Species: Human
    Sample Types: Cell Culture Supernates
  6. The CD40 rs1883832 Polymorphism Affects Sepsis Susceptibility and sCD40L Levels
    Authors: ZL Liu, J Hu, XF Xiao, Y Peng, SP Zhao, XZ Xiao, MS Yang
    Biomed Res Int, 2018;2018(0):7497314.
    Species: Human
    Sample Types: Plasma
  7. Influence of ethnic background on left atrial markers of inflammation, endothelial function and tissue remodelling
    Authors: CD Ruediger, B John, S Kumar, HS Lim, G Rangnekar, KC Roberts-Th, GD Young, D Chase, P Sanders, SR Willoughby
    Indian Pacing Electrophysiol J, 2018;18(1):1-5.
    Species: Human
    Sample Types: Serum
  8. Non-survivor septic patients have persistently higher serum sCD40L levels than survivors
    Authors: L Lorente, MM Martín, A Pérez-Ceja, J Ferreres, J Solé-Violá, L Labarta, C Díaz, A Jiménez
    J Crit Care, 2017;41(0):177-182.
    Species: Human
    Sample Types: Serum
  9. Lipoprotein apheresis in the treatment of dyslipidaemia - the Czech Republic experience
    Authors: V Bláha, M Bláha, M Lánská, D Solichová, L Kujovská K, E Havel, P Vyroubal, Z Zadák, P Žák, L Sobotka
    Physiol Res, 2017;66(0):S91-S100.
    Species: Human
    Sample Types: Serum
  10. Paraoxonase-1 and myeloperoxidase correlate with vascular biomarkers in overweight patients with newly diagnosed untreated hyperlipidaemia
    Authors: A Szentpéter, N Zsíros, VE Varga, H L?rincz, M Katkó, I Seres, P Fülöp, G Paragh, M Harangi
    VASA, 2017;0(0):1-7.
    Species: Human
    Sample Types: Serum
  11. Potential predictive role of chemotherapy-induced changes of soluble CD40 ligand in untreated advanced pancreatic ductal adenocarcinoma
    Onco Targets Ther, 2016;9(0):4681-6.
    Species: Human
    Sample Types: Serum
  12. Endotoxemia contributes to CD27+ memory B-cell apoptosis via enhanced sensitivity to Fas ligation in patients with Cirrhosis
    Sci Rep, 2016;6(0):36862.
    Species: Human
    Sample Types: Plasma
  13. Inhibition of platelet activation and thrombus formation by adenosine and inosine: studies on their relative contribution and molecular modeling.
    Authors: Fuentes E, Pereira J, Mezzano D, Alarcon M, Caballero J, Palomo I
    PLoS ONE, 2014;9(11):e112741.
    Species: Human
    Sample Types: Cell Culture Supernates
  14. Involvement of Rac in thromboxane A2induced human platelet activation: regulation of sCD40 ligand release and PDGFAB secretion.
    Authors: Kageyama Y, Doi T, Matsushima-Nishiwaki R, Iida Y, Akamatsu S, Kondo A, Kuroyanagi G, Yamamoto N, Mizutani J, Otsuka T, Tokuda H, Iida H, Kozawa O, Ogura S
    Mol Med Rep, 2014;10(1):107-12.
    Species: Human
    Sample Types: Plasma
  15. Chlorogenic acid inhibits human platelet activation and thrombus formation.
    Authors: Fuentes E, Caballero J, Alarcon M, Rojas A, Palomo I
    PLoS ONE, 2014;9(3):e90699.
    Species: Human
    Sample Types: Cell Culture Supernates
  16. The evolution of cellular deficiency in GATA2 mutation.
    Authors: Dickinson R, Milne P, Jardine L, Zandi S, Swierczek S, McGovern N, Cookson S, Ferozepurwalla Z, Langridge A, Pagan S, Gennery A, Heiskanen-Kosma T, Hamalainen S, Seppanen M, Helbert M, Tholouli E, Gambineri E, Reykdal S, Gottfreethsson M, Thaventhiran J, Morris E, Hirschfield G, Richter A, Jolles S, Bacon C, Hambleton S, Haniffa M, Bryceson Y, Allen C, Prchal J, Dick J, Bigley V, Collin M
    Blood, 2014;123(6):863-74.
    Species: Human
    Sample Types: Serum
  17. Experimental prestorage filtration removes antibodies and decreases lipids in RBC supernatants mitigating TRALI in vivo.
    Authors: Silliman C, Kelher M, Khan S, LaSarre M, West F, Land K, Mish B, Ceriano L, Sowemimo-Coker S
    Blood, 2014;123(22):3488-95.
    Species: Human
    Sample Types: Serum
  18. Protective mechanisms of guanosine from Solanum lycopersicum on agonist-induced platelet activation: role of sCD40L.
    Authors: Fuentes E, Alarcon M, Astudillo L, Valenzuela C, Gutierrez M, Palomo I
    Molecules, 2013;18(7):8120-35.
    Species: Human
    Sample Types: Platelets
  19. Decreased plasma cytokines are associated with low platelet counts in aplastic anemia and immune thrombocytopenic purpura.
    J. Thromb. Haemost., 2012;10(8):1616-23.
    Species: Human
    Sample Types: Serum
  20. Elderly trauma patients have high circulating noradrenaline levels but attenuated release of adrenaline, platelets, and leukocytes in response to increasing injury severity.
    Authors: Johansson PI, Sorensen AM, Perner A, Welling KL, Wanscher M, Larsen CF, Ostrowski SR
    Crit. Care Med., 2012;40(6):1844-50.
    Species: Human
    Sample Types: Plasma
  21. High sCD40L levels early after trauma are associated with enhanced shock, sympathoadrenal activation, tissue and endothelial damage, coagulopathy and mortality.
    Authors: Johansson PI, Sorensen AM, Perner A, Welling KL, Wanscher M, Larsen CF, Ostrowski SR
    J. Thromb. Haemost., 2011;10(2):207-16.
    Species: Human
    Sample Types: Plasma
  22. Prospective study of circulating soluble CD40 ligand concentrations and the incidence of cardiovascular disease in a nested prospective case-control study of older men and women.
    Authors: Jefferis BJ, Whincup PH, Welsh P, Wannamethee SG, Rumley A, Lawlor DA, Ebrahim S, Lowe GD
    J. Thromb. Haemost., 2011;9(8):1452-9.
    Species: Human
    Sample Types: Serum
  23. Pre-operative platelet-bound CD40 ligand is probably associated with peri-operative cardiac events in hip and knee arthroplasty.
    Authors: Ray MJ, Calabro LJ, Sirisena T, Crawford SA, Crawford RW, Walters DL
    Eur. J. Clin. Invest., 2010;40(6):497-503.
    Species: Human
    Sample Types: Plasma
  24. Matrix metalloproteinase-9 and plasminogen activator inhibitor-1 are associated with right ventricular structure and function: the MESA-RV Study.
    Authors: Kawut SM, Barr RG, Johnson WC
    Biomarkers, 2010;15(8):731-8.
    Species: Human
    Sample Types: Plasma
  25. Validity of multiplex-based assays for cytokine measurements in serum and plasma from "non-diseased" subjects: comparison with ELISA.
    Authors: Dossus L, Becker S, Achaintre D, Kaaks R, Rinaldi S
    J. Immunol. Methods, 2009;350(1):125-32.
    Species: Human
    Sample Types: Plasma
  26. Interleukin-18 is a strong predictor of cardiovascular events in elderly men with the metabolic syndrome: synergistic effect of inflammation and hyperglycemia.
    Authors: Troseid M, Seljeflot I, Hjerkinn EM, Arnesen H
    Diabetes Care, 2009;32(3):486-92.
    Species: Human
    Sample Types: Plasma
  27. Platelet turnover, coagulation factors, and soluble markers of platelet and endothelial activation in essential thrombocythemia: relationship with thrombosis occurrence and JAK2 V617F allele burden.
    Authors: Arellano-Rodrigo E, Alvarez-Larran A, Reverter JC, Colomer D, Villamor N, Bellosillo B, Cervantes F
    Am. J. Hematol., 2009;84(2):102-8.
    Species: Human
    Sample Types: Plasma
  28. A plant stanol yogurt drink alone or combined with a low-dose statin lowers serum triacylglycerol and non-HDL cholesterol in metabolic syndrome patients.
    Authors: Plat J, Brufau G, Dallinga-Thie GM, Dasselaar M, Mensink RP
    J. Nutr., 2009;139(6):1143-9.
    Species: Human
    Sample Types: Plasma
  29. Plasma concentration of SCUBE1, a novel platelet protein, is elevated in patients with acute coronary syndrome and ischemic stroke.
    Authors: Dai DF, Thajeb P, Tu CF, Chiang FT, Chen CH, Yang RB, Chen JJ
    J. Am. Coll. Cardiol., 2008;51(22):2173-80.
    Species: Human
    Sample Types: Plasma
  30. Analysis of platelet alpha2-adrenergic receptor activity in stable coronary artery disease patients on dual antiplatelet therapy.
    Authors: Beres BJ, Toth-Zsamboki E, Vargova K, László A, Masszi T, Kerecsen G, Preda I, Kiss RG
    Thromb. Haemost., 2008;100(5):829-38.
    Species: Human
    Sample Types: Plasma
  31. Soluble CD40 ligand as a biomarker for storage-related preanalytic variations of human serum.
    Authors: Lengelle J, Panopoulos E, Betsou F
    Cytokine, 2008;44(2):275-82.
    Species: Human
    Sample Types: Serum
  32. Fish oil in combination with high or low intakes of linoleic acid lowers plasma triacylglycerols but does not affect other cardiovascular risk markers in healthy men.
    Authors: Damsgaard CT, Frokiaer H, Andersen AD, Lauritzen L
    J. Nutr., 2008;138(6):1061-6.
    Species: Human
    Sample Types: Serum
  33. Hyperglycemia is associated with enhanced thrombin formation, platelet activation, and fibrin clot resistance to lysis in patients with acute coronary syndrome.
    Authors: Undas A, Wiek I, Stepien E, Zmudka K, Tracz W
    Diabetes Care, 2008;31(8):1590-5.
    Species: Human
    Sample Types: Plasma
  34. Rosiglitazone produces a greater reduction in circulating platelet activity compared with gliclazide in patients with type 2 diabetes mellitus--an effect probably mediated by direct platelet PPARgamma activation.
    Authors: Khanolkar MP, Morris RH, Thomas AW, Bolusani H, Roberts AW, Geen J, Jackson SK, Evans LM
    Atherosclerosis, 2007;197(2):718-24.
    Species: Human
    Sample Types: Serum
  35. Myeloid-related protein 8/14 complex is released by monocytes and granulocytes at the site of coronary occlusion: a novel, early, and sensitive marker of acute coronary syndromes.
    Authors: Altwegg LA, Neidhart M, Hersberger M, Muller S, Eberli FR, Corti R, Roffi M, Sutsch G, Gay S, von Eckardstein A, Wischnewsky MB, Luscher TF, Maier W
    Eur. Heart J., 2007;28(8):941-8.
    Species: Human
    Sample Types: Serum
  36. Relationship of soluble CD40 ligand to vascular endothelial growth factor, angiopoietins, and tissue factor in atrial fibrillation: a link among platelet activation, angiogenesis, and thrombosis?
    Authors: Choudhury A, Freestone B, Patel J, Lip GY
    Chest, 2007;132(6):1913-9.
    Species: Human
    Sample Types: Plasma
  37. Multiple biomarker use for detection of adverse events in patients presenting with symptoms suggestive of acute coronary syndrome.
    Authors: Apple FS, Pearce LA, Chung A, Ler R, Murakami MM
    Clin. Chem., 2007;53(5):874-81.
    Species: Human
    Sample Types: Plasma
  38. Relation of improvement in endothelium-dependent flow-mediated vasodilation after rosiglitazone to changes in asymmetric dimethylarginine, endothelin-1, and C-reactive protein in nondiabetic patients with the metabolic syndrome.
    Authors: Wang TD, Chen WJ, Cheng WC, Lin JW, Chen MF, Lee YT
    Am. J. Cardiol., 2006;98(8):1057-62.
    Species: Human
    Sample Types: Plasma
  39. Effects of rosiglitazone alone and in combination with atorvastatin on nontraditional markers of cardiovascular disease in patients with type 2 diabetes mellitus.
    Authors: Chu CS, Lee KT, Lee MY, Su HM, Voon WC, Sheu SH, Lai WT
    Am. J. Cardiol., 2006;97(5):646-50.
    Species: Human
    Sample Types: Serum
  40. Effects of simvastatin on angiogenic growth factors released at the site of microvascular injury.
    Authors: Undas A, Celinska-Lowenhoff M, Stepien E, Nizankowski R, Tracz W, Szczeklik A
    Thromb. Haemost., 2006;95(6):1045-7.
    Species: Human
    Sample Types: Plasma
  41. Humoral and cellular factors responsible for coronary collateral formation.
    Authors: Sherman JA, Hall A, Malenka DJ, De Muinck ED, Simons M
    Am. J. Cardiol., 2006;98(9):1194-7.
    Species: Human
    Sample Types: Plasma
  42. No difference in the effects of clopidogrel and aspirin on inflammatory markers in patients with coronary heart disease.
    Authors: Solheim S, Pettersen AA, Arnesen H, Seljeflot I
    Thromb. Haemost., 2006;96(5):660-4.
    Species: Human
    Sample Types: Plasma


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