Human CD40L ELISA Kit

Catalog No.: BEK1030

Size: 96T

Range: 62.5 pg/ml-4000 pg/ml

Sensitivity < 15 pg/ml

Storage and Expiration: Store at 2-8℃ for 6 months, or at -20℃ for 12 months. 

Application: For quantitative detection of CD40L in human serum, body fluids, tissue lysates or cell culture supernates.

Introduction

CD40L, also called CD154, is a 261 amino acids protein is a member of the TNF superfamily of molecules. It is primarily expressed on activated T cells but is also found in a soluble form. It binds to CD40 on antigen-presenting cells (APC), which leads to many effects depending on the target cell type. In general, CD40L plays the role of a costimulatory molecule and induces activation in APC in association with T cell receptor stimulation by MHC molecules on the APC. In total CD40L has three binding partners: CD40, α5β1 integrin and αIIbβ3.

Principle of the Assay

This kit was based on sandwich enzyme-linked immune-sorbent assay technology. Anti-P- Cadherin polyclonal antibody was pre-coated onto 96-well plates. And the biotin conjugated anti-CD40L polyclonal antibody was used as detection antibodies. The standards, test samples and biotin conjugated detection antibody were added to the wells subsequently, and wash with wash buffer. Avidin-Biotin-Peroxidase Complex was added and unbound conjugates were washed away with wash buffer. TMB substrates were used to visualize HRP enzymatic reaction. TMB was catalyzed by HRP to produce a blue color product that changed into yellow after adding acidic stop solution. The density of yellow is proportional to the CD40L amount of sample captured in plate. Read the O.D. absorbance at 450nm in a microplate reader, and then the concentration of CD40L can be calculated.

Kit components

1. One 96-well plate pre-coated with anti-Human CD40L antibody
2. Lyophilized human CD40L standards: 2 tubes (10 ng / tube)                     
3. Sample / Standard diluent buffer: 30ml
4. Biotin conjugated anti-human CD40L antibody (Concentrated): 130μl. Dilution: 1:100
5. Antibody diluent buffer: 12ml
6. Avidin-Biotin-Peroxidase Complex (ABC) (Concentrated): 130μl. Dilution: 1:100
7. ABC diluent buffer: 12ml
8. TMB substrate: 10ml
9. Stop solution: 10ml
10. Wash buffer (25X): 30ml

Note: Reconstitute standards and test samples with Kit Component 3.

Material Required But Not Provided

1. 37℃ incubator
2. Microplate reader (wavelength: 450nm)
3. Precise pipette and disposable pipette tips
4. Automated plate washer
5. ELISA shaker
6. 1.5ml of Eppendorf tubes
7. Plate cover
8. Absorbent filter papers
9. Plastic or glass container with volume of above 1L