| Field | Specification |
|---|---|
| Target | |
| Alternative names | CD40 ligand;CD40-L;T-cell antigen Gp39;TNF-related activation protein;TRAP;Tumor necrosis factor ligand superfamily member 5;CD154;CD40 ligand, membrane form;CD40 ligand, soluble form;CD40LG;CD40L, TNFSF5, TRAP; |
| Gene ID | |
| UniProt # | |
| Reactivity | |
| Applications | |
| Immunogen | Expression system for standard: E.coli; Immunogen sequence: E108-L261 |
| Expression system | |
| Expression region | |
| Assay type | |
| Sample type(s) | Cell Culture Supernatant, Cell Lysate, Serum, Plasma |
| Sensitivity | |
| Detection range | |
| Assay time | |
| Storage | |
| Catalog no. (Mfr.) | |
| Main SKU |
Assay Principle
This sandwich ELISA quantifies human CD40 ligand (gene CD40LG) in cell culture supernatants, cell lysates, serum and plasma (heparin, EDTA). Strips are pre-coated with a capture antibody. After the sample is added, a biotinylated detection antibody binds the captured protein, and an avidin–biotin–peroxidase complex supplies the enzyme. TMB develops color in proportion to the bound target; the reaction is stopped and read at 450 nm, and concentrations are read from the standard curve.
CD40 ligand (CD40LG, also called CD154) is a TNF family protein expressed mainly on activated CD4+ T cells and platelets. It binds CD40 to drive B-cell activation, immunoglobulin class switching and dendritic cell maturation, and loss of function causes X-linked hyper-IgM syndrome.
Kit Components
| Anti-Human CD40LG Pre-coated 96-well strip microplate | 1 plate (96 wells) |
|---|---|
| Human CD40LG Standard | 2 vials, 10 ng/tube |
| Human CD40LG Biotinylated antibody (100x) | 100 µL |
| Avidin-Biotin-Peroxidase Complex (100x) | 100 µL |
| Sample Diluent | 30 mL |
| Antibody Diluent | 12 mL |
| Avidin-Biotin-Peroxidase Diluent | 12 mL |
| Color Developing Reagent (TMB) | 10 mL |
| Stop Solution | 10 mL |
| Wash Buffer (25x) | 20 mL |
| Adhesive plate sealers | 4 |
Also required (not supplied): microplate reader for 450 nm; incubator; automated plate washer (optional); precision pipettes for 0.5 µL through 1 mL volumes of aqueous solutions; multichannel pipettes (recommended for many samples); deionized or distilled water; 500 mL graduated cylinders; test tubes for dilution.
Kit components are not sold separately.
Performance Data
| Species | Human |
|---|---|
| Sample types | Cell culture supernatants, cell lysates, serum and plasma (heparin, EDTA) |
| Assay range | 62.5 pg/mL–4,000 pg/mL |
| Sensitivity | <15 pg/mL |
| Standard | Recombinant human CD40LG (Glu108–Leu261), expressed in E. coli |
| Cross-reactivity | None detected against related proteins in supplier testing |
| Intra-assay CV | 5.0–7.6% (3 samples, n = 16 each) |
| Inter-assay CV | 6.7–8.6% (3 samples, n = 24 each) |
| Format | 96 wells, removable strips |
Example standard curve (OD450, pg/mL): 0 = 0.063; 62.5 = 0.151; 125 = 0.225; 250 = 0.372; 500 = 0.623; 1000 = 0.981; 2000 = 1.518; 4000 = 2.046. Run a new standard curve on every plate.
Storage & Handling
Store at 4 °C for up to 6 months or −20 °C for up to 12 months. Avoid repeated freeze–thaw cycles. Ships on gel ice and tolerates up to 3 days at room temperature; freeze on arrival.
Safety
Sample data
| Concentration (pg/ml) | 0 | 62.5 | 125 | 250 | 500 | 1000 | 2000 | 4000 |
| O.D. | 0.063 | 0.151 | 0.225 | 0.372 | 0.623 | 0.981 | 1.518 | 2.046 |
Intra/inter assay consistency
| Intra-Assay Precision | Inter-Assay Precision | |||||
|---|---|---|---|---|---|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 16 | 16 | 16 | 24 | 24 | 24 |
| Mean (pg/ml) | 203 | 762 | 1128 | 198 | 762 | 1161 |
| Standard deviation | 15.42 | 58.1%8 | 56.4 | 17.02 | 61.72 | 77.78 |
| CV (%) | 7.6% | 7.4% | 5% | 8.6% | 8.1% | 6.7% |
Kit components
Description|Quantity Pre-coated 96-well strip microplate|1 Standard|2 vials Biotinylated antibody (100x)|100ul Avidin-Biotin-Peroxidase Complex (100x)|100ul Sample Diluent|30ml Antibody Diluent|12ml Avidin-Biotin-Peroxidase Diluent|12ml Color Developing Reagent (TMB)|10ml Stop Solution|10ml Wash Buffer (25x)|20ml Adhesive plate sealers|4Materials required but not provided
- Microplate Reader capable of reading absorbance at 450nm.
- Incubator.
- Automated plate washer (optional).
- Pipettes and pipette tips capable of precisely dispensing 0.5 µl through 1 ml volumes of aqueous solutions.
- Multichannel pipettes are recommended for large amount of samples.
- Deionized or distilled water.
- 500ml graduated cylinders.
- Test tubes for dilution.
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Portillo Jose-Andres C. et al. (2017) CD40 in Retinal Müller Cells Induces P2X7-Dependent Cytokine Expression in Macrophages/Microglia in Diabetic Mice and Development of Early Experimental Diabetic Retinopathy. Diabetes. 10.2337/db16-0051
Alessandra D’Amico et al. (2022) Platelet Activation Favours NOX2-Mediated Muscle Damage in Elite Athletes: The Role of Cocoa-Derived Polyphenols. Nutrients. 10.3390/nu14081558
