Most
fluorescence-based bioanalytical applications need labeling
of analytes. Conventional labeling requires washing to remove the
excess fluorescent labels and reduce the noise signals. These pretreatments
are labor intensive and need specialized equipment, hindering portable
applications in resource-limited areas. Herein, we use the aqueous
two-phase system (ATPS) to realize the partitioning-induced isolation
of labeled analytes from background signals without extra processing
steps. ATPS is formed by mixing two polymers at sufficiently high
concentrations. ATPS-based isolation is driven by intrinsic affinity
differences between analytes and excess labels. To demonstrate the
partitioning-induced isolation and analysis, fluorescein isothiocyanate
(FITC) is selected as the interfering fluorophore, and a monoclonal
antibody (IgG) is used as the analyte. To optimize ATPS compositions,
different molecular weights and mass fractions of polyethylene glycol
(PEG) and dextran and different phosphate-buffered saline (PBS) concentrations
are investigated. Various operational scales of our approach are demonstrated,
suggesting its compatibility with various bioanalytical applications.
In centimeter-scale ATPS, the optimized distribution ratios of IgG
and FITC are 91.682 and 0.998 using PEG 6000 Da and dextran 10,000
Da in 10 mM PBS. In millimeter-scale ATPS, the analyte is enriched
to 6.067 fold using 15 wt % PEG 35,000 Da and 5 wt % dextran 500,000
Da in 10 mM PBS. In microscale ATPS, analyte dilutions are isolated
into picoliter droplets, and the measured fluorescence intensities
linearly correlated with the analyte concentrations (R
2 = 0.982).