Encyclopedia of Experiments: Immunology
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Transcript
Begin by assembling a pre-treated microarray assay slide with distinct spots containing protein-specific capture antibodies for multiple protein detection.
Load the dilutions of the protein standard and the pre-diluted serum samples into separate wells. These dilutions create a range of protein concentrations for accurate quantification.
Incubate to facilitate protein-antibody interactions. Wash to remove unbound proteins, disassemble, and dry the slide.
Invert the assay slide in a snap apparatus containing a rehydrated transfer slide spotted with protein-specific biotinylated detection antibodies in the same positions as the assay slide.
Close the apparatus and press to snap the slides over each other. This facilitates the transfer of the detection antibodies to corresponding spots on the assay slide, minimizing the cross-reactivity.
Separate the slides to allow the interaction of detection antibodies with bound proteins, forming immunocomplexes.
Reassemble the assay slide, rinse it, and introduce fluorophore-conjugated streptavidin, interacting with biotinylated antibodies.
Using a scanner, detect fluorescence spots at various locations, indicating multiple proteins in the serum sample. Compare the spot intensities with the standard for protein quantification.
