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Immunoassays | Oncology | Inflammation & Cytokines

Erenna® Human VEGF Immunoassay Kit (Cat# 03-0068-xx)

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

Assay Performance

TABLE 1. Analytical sensitivity of the Erenna® Human VEGF Immunoassay Kit1
Lower Limit of Detection 0.05 pg/mL
Lower Limit of Quantification2 0.2 pg/mL
Upper Limit of Quantification 100 pg/mL
Low-end CV% Range 2 - 15%
Low-end CV% Average 7%
Recommended Sample Volume 100 μL
Minimum Sample Volume Required3 1 μL

1 See product insert for updated values
2 LLoQ = 20% CV and ± 20% recovery
3 based upon median [VEGF-A (human)] in a healthy reference

FIGURE 1. [VEGF] in EDTA plasma from 10 healthy donors, with median and interquartile range. The Erenna® Human VEGF Immunoassay Kit reliably quantifies VEGF in healthy subjects, who have a median [VEGF] of 66.5 pg/mL that is well above the detection limit of 0.05 pg/mL.

FIGURE 2. Erenna® Human VEGF 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

VEGF-A, commonly known as VEGF, is one member of a family of secreted glycoproteins that promote endothelial cell growth, survival, migration, and vascular permeability, all of which contribute to angiogenesis. The binding of VEGF to its receptor triggers the activation of a cell signaling pathway that is critical for the growth of blood vessels from pre-existing vasculature. VEGF is implicated in a variety of diseases, including several cancers, age-related macular degeneration, diabetic retinopathy and rheumatoid arthritis. With the effi cacy seen with drugs that target VEGF and its pathway, VEGF will remain an attractive candidate for the development of therapies for these diseases, particularly cancer.

TABLE 2. Additional UniProtKB/Swiss-Prot Information*
Protein Name: Vascular endothelial growth factor A
UniProtKB/Swiss-Prot ID: P15692
Protein-protein interaction database: P15692
Alternative Names: Vascular permeability factor
Gene Names: VEGFA
Synonym Gene Names: VEGF
Function: Growth factor active in angiogenesis, vasculogenesis and endothelial cell growth. Induces endothelial cell proliferation, promotes cell migration, inhibits apoptosis and induces permeabilization of blood vessels. Binds to the FLT1/VEGFR1 and KDR/VEGFR2 receptors, heparan sulfate and heparin. NRP1/Neuropilin-1 binds isoforms VEGF-165 and VEGF-145. Isoform VEGF165B binds to KDR but does not activate downstream signaling pathways, does not activate angiogenesis and inhibits tumor growth.
Biological Processes: angiogenesis, differentiation, positive regulation of mast cell chemotaxis
Molecular Function: chemoattractant activity, extracellular matrix binding, cytokine activity, vascular endothelial growth factor receptor binding

* Source Info from www.uniprot.org.

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