Encyclopedia of Experiments: Immunology
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Transcript
To detect anti-aquaporin-4 immunoglobulin-G or anti-AQP4 IgG — a biomarker of an autoimmune disorder, begin with a multi-well reagent tray.
Fill the wells individually with a positive control containing anti-AQP4 IgGs and pre-diluted serum samples.
Position a biochip with reaction fields over the reagent tray, ensuring direct contact.
Each biochip field contains two subsections, one coated with transfected cells expressing aquaporin-4 — a water-channel protein, and the other with untransfected cells lacking aquaporin-4.
The anti-AQP4 IgGs from the samples bind to the aquaporin-4 channels on transfected cells.
Remove the biochip and rinse with a detergent-supplemented wash buffer to eliminate non-specific interactions.
Overlay with fluorophore-labeled-secondary antibodies, interacting with pre-bound anti-AQP4 IgGs.
Wash, add a mounting medium, and observe under a fluorescence microscope.
The serum sample shows stronger fluorescence in the transfected subsections compared to the untransfected ones.
This fluorescence pattern aligns with that of the positive control, confirming the presence of anti-AQP4 IgGs in the serum.
