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Immunoassays | Inflammation & Cytokines

Erenna® IL-5 Immunoassay Kit (Cat# 03-0053-xx)

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The Erenna® IL-5 Immunoassay Kit uses a quantitative fluorescent sandwich immunoassay technique to measure IL-5 in plasma and serum samples. A capture antibody specific for IL-5 has been pre-coated onto paramagnetic microparticles (MP). The user pipettes MP, standards, and samples into uncoated microplate wells. During incubation, the IL-5 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 IL-5 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 IL-5 present in the sample when captured. The amount of IL-5 in unknown samples is interpolated from a standard curve.

Assay Performance

TABLE 1. Analytical sensitivity of the Erenna® IL-5 Immunoassay Kit1
Lower Limit of Detection 0.17 pg/mL
Lower Limit of Quantification2 3.91 pg/mL
Upper Limit of Quantification 500 pg/mL
Low-end CV% Range 4 - 13%
Low-end CV% Average 9%
Recommended Sample Volume 100 µL
Minimum Sample Volume Required3 100 μL

1 See product insert for updated values
2 LLoQ = 20% CV and ± 20% recovery
3 based upon median [IL-5] in a healthy reference

FIGURE 1. [IL-5] in EDTA plasma from 10 healthy donors (7 samples < LLoQ), with median and interquartile range. Erenna® IL-5 Immunoassay Kit reliably quantifies IL-5 in EDTA plasma from healthy subjects, who have a median [IL-5] of 4.52 pg/mL that is well above the detection limit of 0.17 pg/mL.

FIGURE 2. Erenna® IL-5 Immunoassay Kit low-end standard curve signal (left) and curve fit (right).

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

Biology and Disease

Interleukin-5 (IL-5) is a cytokine secreted mainly by T-cells and mast cells upon stimulation with antigens and allergens. It has pleiotropic effects toward B-cells and is involved in the production of immunoglobulins. More importantly, IL-5 acts in synergy with GM-CSF to govern the proliferation and maturation of eosinophils. A pronounced level of eosinophils is commonly associated with the development of allergic conditions such as asthma, eosinophilic esophagitis and hypereosinophilic syndrome. As such, IL-5 is highly regarded as a crucial mediator of the inflammatory pathway concerning allergic reactions. Current studies suggest that IL-5 blockade is a promising intervention for the treatment of allergic diseases.

TABLE 2. Additional UniProtKB/Swiss-Prot Information*
Protein Name: Interleukin-5
UniProtKB/Swiss-Prot ID: P05113
Protein-protein interaction database: P05113
Alternative Names: B-cell differentiation factor I, Eosinophil differentiation factor, T-cell replacing factor
Gene Names: IL5
Synonym Gene Names:
Function: Factor that induces terminal differentiation of late-developing B-cells to immunoglobulin secreting cells.
Biological Processes: immune response, inflammatory response, positive regulation of peptidyl-tyrosine phosphorylation
Molecular Function: cytokine activity, growth factor activity, interleukin-5 receptor binding

* Source Info from www.uniprot.org.

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