Human VEGF Quantikine QuicKit ELISA

Catalog #: QK293
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QK293

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QC285: Human VEGF Quantikine QuicKit Control Group 285

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All VEGF ELISAs

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Summary Sample Values Recovery Linearity Data Examples Product Datasheets Preparation and Storage Background Related Research Areas

Human VEGF Quantikine QuicKit ELISA Summary

Assay Length

80 minutes

Sample Type & Volume Required Per Well

Cell Culture Supernates (50 uL), Serum (50 uL), EDTA Plasma (50 uL), Heparin Plasma (50 uL), Saliva (25 uL), Urine (13 uL), Human Milk (10 uL)

Sensitivity

3.19 pg/mL

Assay Range

28.1 - 1,800 pg/mL (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma, Saliva, Urine, Human Milk)

Specificity

Natural and recombinant human VEGF.

Cross-reactivity

< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.

Interference

No significant interference observed with available related molecules.

Sample Values

Serum/Plasma/Saliva/Urine/Human Milk - Samples from apparently healthy volunteers were evaluated for the presence of human VEGF in this assay. No medical histories were available for the donors used in this study.

Sample Type	Mean (pg/mL)	Range (pg/mL)	Standard Deviation (pg/mL)
Serum (n=10)	519	175-1093	352
EDTA plasma (n=10)	79.4	40.6-183	43.1
Heparin plasma (n=10)	69.8	38.9-105	23.7
Saliva (n=4)	3521	2055-4880	1374
Urine (n=4)	566	219-1184	429
Human milk (n=4)	27,535	23,100-31,888	4620

Cell Culture Supernates:

PBMCs were isolated from a single donor over a Ficoll-Paque PLUS density gradient. PBMCs were seeded at 1 x 106 /mL and cultured in RPMI 1640 supplemented with 10% fetal bovine serum, 2 mM L-glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin sulfate. The cells were stimulated with 10 μg/mL PHA for 5 days. Aliquots of the cell culture supernatants were removed, assayed for human VEGF, and measured 169 pg/mL.

JEG-3 human epithelial choriocarcinoma cells were cultured in MEM Earle's Salts and supplemented with 10% FBS. Cells were stimulated with 100 ng/mL of LPS for 24 hours. An aliquot of the cell culture supernate was removed, assayed for human VEGF, and measured 11,100 pg/mL.

THP-1 acute monocytic leukemia cells were cultured in RPMI and supplemented with 10% fetal bovine serum. Cells were stimulated with 1 μg/mL Retionic Acid and 100 ng/mL PMA for 4 days. An aliquot of the cell culture supernate was removed, assayed for human VEGF, and measured 1829 pg/mL.

PC-3 human prostate cancer cells were cultured in RPMI and supplemented with 10% fetal bovine serum until confluent. An aliquot of the cell culture supernate was removed, assayed for human VEGF, and measured 2366 pg/mL.

Product Summary

The Quantikine™ QuicKit™ Human VEGF Immunoassay is a one step, 80-minute solid phase ELISA designed to measure human VEGF levels in cell culture supernates, serum, plasma, saliva, urine, and human milk. It contains Sf 21-expressed recombinant human VEGF and antibodies raised against the recombinant protein. Results obtained using natural human VEGF showed linear curves that were parallel to the standard curves obtained using the QuicKit standards. These results indicate that this kit can be used to determine relative mass values for natural human VEGF.

Recovery

The recovery of human VEGF spiked to three levels in samples throughout the range of the assay was evaluated.

Sample Type	Average % Recovery	Range %
Cell Culture Media (n=4)	99	94-104
EDTA Plasma (n=2)	83	79-88
Heparin Plasma (n=2)	77	70-81
Human Milk (n=2)	102	98-105
Saliva (n=2)	104	97-115
Serum (n=2)	79	74-84
Urine (n=2)	78	70-85

Linearity

To assess the linearity of the assay, samples containing and/or spiked with high concentrations of human VEGF were diluted with calibrator diluent to produce samples with values within the dynamic range of the assay.

Scientific Data

N/A

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Human VEGF Standard Curve

ELISA

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Human VEGF QuicKit ELISA Spiked Linearity Competitor Comparison Samples containing VEGF were diluted with appropriate Calibrator Diluent to produce samples with values within the dynamic range of the assay. The linearity is between 99%-107% compared to 100%-145% for the top competitor.

Product Datasheets

Product Datasheet

COA

SDS

Preparation and Storage

Shipping

The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.

Storage

Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: VEGF

Vascular endothelial growth factor (VEGF or VEGF-A), also known as vascular permeability factor (VPF), is a potent mediator of both angiogenesis and vasculogenesis in the fetus and adult (1-3). It is a member of the PDGF family that is characterized by the presence of eight conserved cysteine residues in a cystine knot structure and the formation of antiparallel disulfide-linked dimers (4). Humans express alternately spliced isoforms of 121, 145, 165, 183, 189, and 206 amino acids (aa) in length (4). VEGF₁₆₅ appears to be the most abundant and potent isoform, followed by VEGF₁₂₁ and VEGF₁₈₉ (3, 4). Isoforms other than VEGF₁₂₁ contain basic heparin-binding regions and are not freely diffusible (4). Human VEGF₁₆₅ shares 88% aa sequence identity with corresponding regions of mouse and rat VEGF. VEGF is expressed in multiple cells and tissues including skeletal and cardiac muscle (5, 6), hepatocytes (7), osteoblasts (8), neutrophils (9), macrophages (10), keratinocytes (11), brown adipose tissue (12), CD34⁺ stem cells (13), endothelial cells (14), fibroblasts, and vascular smooth muscle cells (15). VEGF expression is induced by hypoxia and cytokines such as IL-1, IL-6, IL-8, oncostatin M, and TNF-alpha (3, 4, 9, 16). VEGF isoforms are differentially expressed during development and in the adult (3).

VEGF dimers bind to two related receptor tyrosine kinases, VEGF R1 (also called Flt-1) and VEGF R2 (Flk-1/KDR), and induce their homodimerization and autophosphorylation (3, 4, 7, 17, 18). These receptors have seven extracellular immunoglobulin-like domains and an intracellular split tyrosine kinase domain. They are expressed on vascular endothelial cells and a range of non-endothelial cells. Although VEGF affinity is highest for binding to VEGF R1, VEGF R2 appears to be the primary mediator of VEGF angiogenic activity (3, 4). VEGF165 also binds the semaphorin receptor, neuropilin-1, which promotes complex formation with VEGF R2 (19).

VEGF is best known for its role in vasculogenesis. During embryogenesis, VEGF regulates the proliferation, migration, and survival of endothelial cells (3, 4), thus regulating blood vessel density and size, but playing no role in determining vascular patterns. VEGF promotes bone formation through osteoblast and chondroblast recruitment and is also a monocyte chemoattractant (20-22). After birth, VEGF maintains endothelial cell integrity and is a potent mitogen for micro- and macro-vascular endothelial cells. In adults, VEGF functions mainly in wound healing and the female reproductive cycle (3). In diseased tissues, VEGF promotes vascular permeability. It is thus thought to contribute to tumor metastasis by promoting both extravasation and tumor angiogenesis (23, 24). Various strategies have been employed therapeutically to antagonize VEGF-mediated tumor angiogenesis (25). Circulating VEGF levels correlate with disease activity in autoimmune diseases such as rheumatoid arthritis, multiple sclerosis and systemic lupus erythematosus (26).

Long Name:

Vascular Endothelial Growth Factor

Entrez Gene IDs:

7422 (Human); 22339 (Mouse); 83785 (Rat); 281572 (Bovine); 403802 (Canine); 493845 (Feline); 30682 (Zebrafish)

Alternate Names:

MVCD1; VAS; vascular endothelial growth factor A; Vascular permeability factor; Vasculotropin; VEGF; VEGFA; VEGF-A; VEGFMGC70609; VPF; VPFvascular endothelial growth factor

Related Research Areas

Citation for Human VEGF Quantikine QuicKit ELISA

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

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