Acoustofluidics-manipulated triple-emission fluorescent nanoprobe aggregates with multicolor-variation for colorimetric quantitative assay

[Display omitted] •The b-SiCDs covalently bind in SiO2 NPs to exhibit strong internal standard robustness.•The triple-emission probe is constructed by two ratiometric fluorescent composite NPs.•The probe shows more profuse and wider color variations using two-step response strategy.•Sensitivity for colorimetric assay of Hb is further improved by acoustofluidic technology.•RGB values got by chromaticity analysis APP are used for accurate Hb quantification. Acoustofluidics-assisted colorimetric method, a promising point-of-care test (POCT) strategy, has been applied in convenient detection of target molecules using the sensing nanomaterials, but remains a formidable challenge to design fluorescent probe with profuse color evolution, realizing sensitive, simple and accurate determination of analyte. Here, a novel triple-emission fluorescent probe is constructed through a simple blend of two ratiometric fluorescent probes (blue-organosilane-polymerized carbon dots@SiO2 nanoparticles@red-CdTe/CdS quantum dots (b@SiO2@r) and blue-organosilane-polymerized carbon dots@SiO2 nanoparticles@green-CdTe/CdS quantum dots (b@SiO2@g)) at the optimized volume ratio of 11:8. As signal reporters of the triple-emission probe, outer modified “r” and “g” will be quenched in sequence by target via two-step response strategy while the inner “b” as internal standard remains constant. When acoustic field is applied, the effect of acoustofluidics-based nanoparticles concentration makes triple-emission probe aggregate completely, accompanying fluorescence color signal being enhanced, sensitivity of colorimetric assay being improved and multicolor-variation (from orange to dark-goldenrod to dark-olive-green to sea-green to dark-cyan to final steel-blue) with the increase of analyte amount. For POCT demonstration, hemoglobin (Hb) is used as a model target and the captured fluorescence photos of triple-emission probe aggregates are analyzed by a chromaticity analysis application on the smartphone to calculate the red, green and blue values, which are used for accurate Hb quantification with the limit of detection of 1.99 mg/L. Whole test is finished within ∼12 min. The acoustofluidics-manipulated triple-emission probe biosensing strategy has also been applied to the rapid and visual quantitative detection of Hb in human blood.