PCATALOG
AlEgens Barcodes Combined with AlEgens Nanobeads for High-sensitivity Multiplexed Detection
Pubdate：2021-12-23 Click：907
Suspension arrays based on optical encoded microspheres have attracted great attention for multiplexed detection in gene analysis, protein profiling, early disease diagnosis, treatment monitoring and so on. However, the fluorescence stability of barcodes and detection sensitivity require further improvement to meet the increasing demands of "precision diagnosis".
Methods: This work reports a novel suspension array platform based on extremely stable Ale gens (AIE33 and AIE NIR800) microbeads as barcodes and AlEgens (1,1,2,3,4,5-Hexaphenyl-1H-silole, HPS) nanobeads as fluorescent signal reporter coupled with flow cytometry for multiplexed detection.
Results: Due to the excellent fluorescent signal amplification effect of the HPS nanobeads, our multiplex assay showed enhanced detection sensitivity, compared to multiplex assay using QDs nanobeads (up to 3-fold improvement) and commercial organic dye of phycoerythrin (up to 5-fold improvement) as the fluorescent signal reporters.
Conclusion: Furthermore, validating experiments showed similar detection performance to the clinical gold-standard method of ImmunoCAP for allergen detection in patient serum samples, demonstrating the suspension array platform based on AlEgens microbeads with excellent fluorescence stability and AlEgens nanobeads with strong signal amplification ability is promising for high-sensitivity multiplexed bioassay applications.
Key words: Aggregation-induced emission (AIE); AlEgens barcodes; AIEgens nanobeads;multiplexed detection, allergens
Conclusions
In summary, we introduced AIEgens to suspension arrays for the first time, and successfully constructed a novel suspension array platform based on AIEgens microbeads as barcodes and AIEgens nanobeads as fluorescent signal reporters together with flow cytometry excitation by a single laser for multiplexed detection. We synthesized both AIEgens barcodes and AIEgens nanobeads using the facile Shirasu porous glass (SPG) membrane emulsification method by simply altering the pore diameter of the SPG membrane. The as-prepared beads had a narrow size distribution, good fluorescence uniformity inside beads and excellent fluorescence stability. Their fluorescent intensities were also extremely stable even
in both highly acidic and alkaline solutions, demonstrating superior fluorescence stability among reported optical encoded beads. We successfully produced a 2D AIEgens barcode library containing 30 barcodes based on the ‘single wavelength’ encoding method by varying the emission wavelength and PL intensity levels of AIEgens (HPS, AIE41, AIE33 and AIE NIR800). Moreover, we combined AIE33 and AIE NIR800 microbeads barcodes and HPS nanobeads fluorescent signal reporters with flow cytometer to detect five common allergens in a single sample. The multiplex assay using AIEgens nanobeads as fluorescent signal reporters had higher detection sensitivity, compared with multiplex assays using commercial organic label of phycoerythrin (up to 5-fold improvement) and QDs nanobeads (up to 3-fold improvement) as reporters, indicating the excellent fluorescent signal amplification effect of HPS nanobeads. The results obtained by detecting 95 patient samples exhibit that each allergen achieved similar detection properties in terms of sensitivity and specificity compared with the current clinical gold-standard method of ImmunoCAP. Together the results show that the suspension array platform based on AIEgens microbeads with excellent fluorescence stability and AIEgens nanobeads with strong signal amplification ability for multiplex immunoassays hold promise for biological applications, and can be applied to other proteins, nucleic acids and cytokines in the future.
