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Immunoassays | Inflammation & Cytokines

Erenna® GM-CSF Immunoassay Kit (Cat# 03-0067-xx)

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

Assay Performance

TABLE 1. Analytical sensitivity of the Erenna® GM-CSF Immunoassay Kit1
Lower Limit of Detection 0.004 pg/mL
Lower Limit of Quantification2 0.02 pg/mL
Upper Limit of Quantification 5 pg/mL
Low-end CV% Range 1 - 11%
Low-end CV% Average 6%
Recommended Sample Volume 100 μL
Minimum Sample Volume Required3 50 μL

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

FIGURE 1. [GM-CSF] in EDTA plasma from 10 healthy donors, with median and interquartile range. The Erenna® GM-CSF Immunoassay Kit reliably quantifies GM-CSF in healthy subjects, who have a median [GM-CSF] of 0.2 pg/mL that is well above the detection limit of 0.004 pg/mL.

FIGURE 2. Erenna® GM-CSF 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

Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF) is a cytokine produced by T cells, B cells, macrophages, mast cells and fibroblasts in response to inflammation or infection. Its primary role is to stimulate granulocyte and monocyte production in the bone marrow. Once GM-CSF has bound to its receptor, the activated complex propagates a signaling cascade that increases macrophage counts in order to fight infection. Thus increased GM-CSF can be observed during episodes of infectious disease and in a spectrum of inflammatory disorders such as Crohn’s disease. Conversely, a lack of GM-CSF resulting from autoantibody production has been linked to pulmonary alveolar proteinosis, leukemia, and neutropenia.

TABLE 2. Additional UniProtKB/Swiss-Prot Information*
Protein Name: Granulocyte-macrophage colony-stimulating factor
UniProtKB/Swiss-Prot ID: P04141
Protein-protein interaction database: P04141
Alternative Names: Colony-stimulating factor
Gene Names: CSF2
Synonym Gene Names: GMCSF
Function: Cytokine that stimulates the growth and differentiation of hematopoietic precursor cells from various lineages, including granulocytes, macrophages, eosinophils and erythrocytes.
Biological Processes: immune response, positive regulation of macrophage derived foam cell differentiation, positive regulation of DNA replication
Molecular Function: cytokine activity, growth factor activity, granulocyte macrophage colony-stimulating factor receptor binding

* Source Info from www.uniprot.org.

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