Description
The antibodies in this kit recognize VEGF-A (VEGF165 or isoform d), VEGF162, VEGF145 (isoform e) and VEGF121 (isoform f). One of the sandwiching antibodies is a mouse monoclonal, clone 26503. The other sandwiching antibody is a goat polyclonal raised against human recombinant VEGF165.
Formats:
Our 500 assay point kit allows you to run 500 wells in 96-well or 384-well format, using a 50 µL reaction volume (5 µL of sample).
Our 5,000 assay point kit allows you to run 5,000 wells in 96-well or 384-well format, using a 50 µL reaction volume (5 µL of sample).
Features:
No-wash steps, no separation steps
ELISA alternative technology
Sensitive detection
Broad sample compatibility
Small sample volume
Results in less than 3 hours
Half the time of an ELISA assay
Vascular endothelial growth factor (VEGF-A or VEGF) is a homodimeric 34-42 kDa heparin-binding glycoprotein specific for endothelial cells. Currently at least eight variants of VEGF generated by alternative splicing of the same gene have been identified. The various isoforms have different heparin- and neuropilin(-1 or -2)-binding properties as well as solubility characteristics reflecting the different functional properties of the VEGF forms. VEGF is believed to play important roles in inflammation and during normal and pathological angiogenesis, a process that is associated with wound healing, embryonic development, and growth and metastasis of solid tumors. Elevated levels of VEGF have been observed in synovial fluids of rheumatoid arthritis patients and sera of cancer patients.
AlphaLISA technology allows the detection of molecules of interest in a no-wash, highly sensitive, quantitative assay. In an AlphaLISA assay, a biotinylated anti-analyte antibody binds to the Streptavidin-coated Donor beads while another anti-analyte antibody is conjugated to AlphaLISA Acceptor beads. In the presence of the analyte, the beads come into close proximity. The excitation of the Donor beads causes the release of singlet oxygen molecules that triggers a cascade of energy transfer in the Acceptor beads, resulting in a sharp peak of light emission at 615 nm.
Principle of the AlphaLISA assay
The AlphaLISA assay is based on an AlphaLISA sandwich immunoassay involving a biotinylated anti-analyte antibody bound to Streptavidin-coated AlphaLISA Donor beads and an anti-analyte antibody conjugated to AlphaLISA Acceptor beads. Both antibodies are directed against the analyte of interest. In the presence of the target, both antibodies bind to analyte and the beads come into proximity. The excitation of the Donor beads provokes the release of singlet oxygen molecules that triggers a cascade of energy transfer within the Acceptor beads, resulting in emission with λmax at 615 nm. The intensity of the signal is directly proportional to the concentration of analyte present in the sample.
Protocol of the AlphaLISA assay
The AlphaLISA assay can be run in a 96- or 384-well detection plate (50 µL final). As described here, samples or standards are dispensed directly into the assay plate for the detection of the analyte of interest by AlphaLISA reagents. No washing steps are needed. The protocol can be further miniaturized or upscaled by simply resizing each addition volume proportionally.
