<br/>One-pot processing assays can reduce tedious laboratory work and requires little expertise. Hence, they are more suitable to be integrated into point-of-care devices for large-scale public health screening. However, washing steps are inevitable in many common immunoassays mainly for the addition of excess biomarkers and removal of the unbound ones. Therefore, to combine traditional assays with droplets, cumbersome pretreatment of the sample is required before microfluidic compartmentalization. Here we develop a wash-free, selective affinity-based method to localize biomolecules. The fluorescence-labeled analytes can be spontaneously isolated from excess fluorescent markers inside digital droplets. Taking advantage of different affinities of biomolecules in the aqueous two-phase system (ATPS), this method allows separation of the marked and unmarked components and isolate them into separate areas with different signal intensities. The fluorescence contrast between the core and shell phases inside the monodisperse ATPS double-emulsion droplets indicates whether the target molecules exist, and the intensity difference correlates with analyte loads. The analyte loads measured by fluorescence contrast showed good consistency with fluorescence-labeled analytes processed by standard washing steps. In summary, we demonstrate the reliability of this assay compared to the conventional method and highlight its feature of the entirely wash-free process.

Selective partitioning of biomolecules in all-aqueous double emulsions for isolation and detection of analytes

ACS Spring Meeting 2021

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