Competitive ELISA, also known as inhibition ELISA or competitive immunoassay, is a widely used technique for quantifying antigen levels in a sample. Unlike other ELISA formats, this method relies on the competition between the sample antigen and a labeled reference antigen for binding to a limited quantity of antibodies. As a result, the detection signal is inversely proportional to the antigen concentration in the sample -higher antigen levels result in lower signal intensity. This inverse relationship is particularly advantageous when detecting small molecules or antigens with few available epitopes.
Our competitive ELISA kits are particularly useful in scenarios where direct detection methods are impractical. For example, measuring low-molecular-weight compounds such as hormones, drugs, and metabolites can be challenging using a sandwich ELISA due to the limited number of epitopes available for antibody binding. Our competitive ELISA also excels when using samples that have complex matrices, which may interfere with direct detection, and in cases where the target antigen lacks the two binding sites necessary for a sandwich format. Its high specificity and sensitivity make it a preferred choice for quantifying low-abundance proteins and other analytes with structural constraints.
This competitive ELISA test can be performed using different assay orientations. In the antigen-down format, a reference antigen is pre-coated on the multi-well plate, and the sample containing the antigen of interest is added along with a labeled antibody. In this setup, a higher concentration of sample antigen results in fewer free antibodies available to bind the reference antigen, leading to a lower signal. Alternatively, in the antibody-down format, the primary antibody is immobilized on the plate, and both the labeled antigen and the sample antigen compete for binding. Here, a lower antigen concentration in the sample allows for more labeled antigens to bind, producing a stronger signal.
Our competitive ELISA kits are a versatile and sensitive option for analyzing small molecules, low-abundance proteins, and analytes within complex matrices. Their unique format and flexibility in design make them invaluable tools in research and diagnostic laboratories.
Features
- Our competitive ELISA kits are designed to effectively detect analytes with limited epitopes, making them well-suited for small molecules and short peptides.
- They utilize only one antibody in the assay, reducing potential interference from complex sample matrices.
Applications
- Our competitive ELISA kits are effectively employed for the quantitative measurement of various in serum, EDTA plasma, urine, citrate plasma, heparin plasma, and cell culture media.
- These can also be used for the quantitative measurement of intracellular targets in cell lysates and tissue extract samples.
- Our competitive ELISA kits can also be used for the sensitive and specific detection of other compounds and molecules, such as antibiotics in cell and tissue extract samples from various species, as well as milk and milk powder.
- Our kits enable sensitive, quantitative measurement of proteins with various tags.
What is competitive ELISA, and how does it work?
Competitive ELISA is an immunoassay that quantifies antigen concentration by using a competitive binding approach. In the assay, a fixed amount of labeled antigen and the sample antigen compete for binding to a limited amount of antibody. As the concentration of the sample antigen increases, fewer labeled antigens bind, resulting in a lower detection signal. This inverse relationship between signal intensity and antigen concentration enables precise quantification, making competitive ELISA ideal for small molecules or analytes with limited epitopes.
What factors affect the sensitivity and specificity of competitive ELISA?
Factors affecting the sensitivity and specificity of competitive ELISA include the quality and affinity of the antibodies used, as high-affinity antibodies ensure precise competitive binding. The concentration and labeling efficiency of both the sample and reference antigens play a critical role in achieving optimal competition. Assay conditions, such as incubation time, temperature, and effective blocking, help minimize non-specific binding and reduce background noise. Additionally, the performance of the detection system and the handling of sample matrix effects further influence the assay’s overall sensitivity and specificity.
What types of samples can be used in a competitive ELISA experiment?
Competitive ELISA is a versatile assay that can be applied to a wide range of sample types, including serum, various plasma types (EDTA, citrate, heparin), and urine. It is also suitable for use with cell culture media, tissue extracts, and cell extracts, making it ideal for a range of experimental applications.
What are the advantages of using competitive ELISA over other immunoassay techniques?
Competitive ELISA is particularly advantageous for detecting small molecules or analytes with a single epitope, where sandwich assays are not feasible. It requires only one specific antibody, which simplifies assay design and can reduce costs. Additionally, this format minimizes matrix effects and non-specific binding, providing high sensitivity and specificity even in complex samples.