Short CommunicationsFIDA automation yields sub-femtomolar limit of detection for Aβ aggregates in body fluids

- sFIDA assay was successfully implemented on a fully automated robot platform.
- Aβ silica nanoparticles were spiked into CSF/PBS to investigate the assay outcome.
- sFIDA assay automation results were linear over four orders of magnitude.
- sFIDA assay automation resulted in highly enhanced intra-assay precision.
- Sensitivity was enhanced to a limit of detection in the sub-femtomolar range.

ObjectivesAlzheimer's disease (AD) is a neurodegenerative disorder with yet non-existent therapeutic and limited diagnostic options. Reliable biomarker-based AD diagnostics are of utmost importance for the development and application of therapeutic substances. We have previously introduced a platform technology designated 'sFIDA' for the quantitation of amyloid β peptide (Aβ) aggregates as AD biomarker. In this study we implemented the sFIDA assay on an automated platform to enhance robustness and performance of the assay.Design and methodsIn sFIDA (surface-based fluorescence intensity distribution analysis) Aβ species are immobilized by a capture antibody to a glass surface. Aβ aggregates are then multiply loaded with fluorescent antibodies and quantitated by high resolution fluorescence microscopy. As a model system for Aβ aggregates, we used Aβ-conjugated silica nanoparticles (Aβ-SiNaPs) diluted in PBS buffer and cerebrospinal fluid, respectively. Automation of the assay was realized on a liquid handling system in combination with a microplate washer.ResultsThe automation of the sFIDA assay results in improved intra-assay precision, linearity and sensitivity in comparison to the manual application, and achieved a limit of detection in the sub-femtomolar range.ConclusionsAutomation improves the precision and sensitivity of the sFIDA assay, which is a prerequisite for high-throughput measurements and future application of the technology in routine AD diagnostics.

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