Encyclopedia of Experiments: Immunology
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Transcript
To detect a disease-associated protein biomarker, take an immunoassay plate.
The plate contains pre-filled separation and stacking matrices required for size-based protein separation.
Load the proteins and immunoassay reagents into distinct plate wells.
Initiate the run inside a CEI analyzer. The capillary cartridge in the analyzer aspirates the matrices to form a gel within each capillary.
The capillary then aspirates the proteins. During electrophoresis, the proteins migrate and separate based on their molecular weights.
The analyzer introduces UV light, immobilizing the proteins in the capillary.
The wash buffer removes the matrices and unbound proteins. The blocking buffer then flows in, blocking non-specific interaction sites.
The analyzer introduces primary antibodies that bind to the biomarker, followed by peroxidase-conjugated secondary antibodies interacting with the primary antibodies.
Wash and then add the chemiluminescence substrate. Peroxidase oxidizes the substrate, emitting luminescence.
Measure the luminescence intensity and compare it against the protein standard for biomarker detection.
