Immunoassays | Prototype

Erenna® Mouse TNFα Immunoassay Prototype Assay (Cat# 03-0060-AA)

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Introduction

The Erenna® Mouse TNFα Immunoassay Evaluation Reagent Kit uses a quantitative fluorescent sandwich immunoassay technique to measure TNFα in plasma and serum samples. A capture antibody specific for TNFα has been pre-coated onto paramagnetic microparticles (MP). The user pipettes MP, standards, and samples into uncoated microplate wells. During incubation, the TNFα present in the sample binds to the capture antibody on the coated MP. Unbound molecules are washed away during the subsequent buffer exchange and wash steps. Fluor-labeled detection antibody is added to each well and incubated. This detection antibody recognizes and binds to TNFα that has been captured onto the MP. During the following wash step the MPs are transferred to a clean plate. Elution buffer is then added and incubated. The elution buffer dissociates the bound protein sandwich from the MP surface, releasing the labeled antibodies. These antibodies are separated during transfer to a final microplate. The plate is loaded into the Erenna System where the labeled molecules are detected and counted. The number of fluor-labeled detection antibodies counted is directly proportional to the amount of TNFα present in the sample when captured. The amount of TNFα in unknown samples is interpolated from a standard curve.

Assay Performance

¹ See product insert for updated values
² LLoQ = 20% CV and ± 20% recovery

FIGURE 2. Erenna® Mouse TNFα Immunoassay Evaluation Reagent Kit low-end standard curve signal (left) and curve fit (right). The Erenna® Mouse TNFα Immunoassay Evaluation Reagent Kit has a detection limit of 0.08 pg/mL, providing accurate quantification of mTNFα in only 20 μL of plasma. This assay enables better preclinical study design and long-term monitoring of individual mice over time.

Representative data shown for demonstration purposes only. Individual results may vary depending upon samples tested and protocol used.

Biology and Disease

Tumor necrosis factor alpha (TNFα) is a pro-inflammatory cytokine with cytotoxic activity that is primarily produced by macrophages. TNFα is involved in systemic inflammation and is a member of the group of cytokines that stimulate the acute phase reaction. When administered to animals or humans it causes inflammation, fever, cardiovascular eff ects, hemorrhage, coagulation, and acute phase responses similar to those seen during acute infections and shock states. By promoting the inflammatory response, TNFα is associated with many autoimmune disorders such as rheumatoid arthritis, ankylosing spondylitis, Crohn’s disease, psoriasis, and refractory asthma. TNFα also plays a role in cancer and regulates apoptotic cell death, cellular proliferation, tumorigenesis, and viral replication.

* Source Info from www.uniprot.org.

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