The construction of multiple fluorescent nanosensors for intracellular studies is a challenging task because spectral overlap of indicator probes can lead to cross-talk and mutual interference. This work describes biodegradable nanosensors that can simultaneously measure three intracellular parameters (temperature, pH and oxygen concentration). Bovine serum albumin (BSA) is selected as the scaffold to construct the triple nanosensor by covalent immobilization four fluorophores on BSA. The following luminophores were used: (a) fluorescein as a probe for pH values, (b) a platinum(II) porphyrin complex for oxygen; (c) a europium(III) clathrate complex for temperature, and (d) a rhodamine B as a reference dye. The nanoparticles have a size of 20 nm and show excellent biocompatibility and good brightness. The nanosensors were used for ratiometric imaging of intracellular pH values, oxygen and temperature in HeLa cells. The triple nanosensor responds reversibly and this can be used for real-time tracing of these key parameters. Owing to their biodegradable feature, the use of this kind of triple nanosensor reduce the stress on cellular activities because less nanosensors can be used to gather the total information.
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This work was financially supported by the National Natural Science Foundation of China (21775029), the National Key R&D Program of China (2017YFC0906800), the Recruitment Program of Global Experts (1000 Talent program) in China, and the Program for Professor of Special Appointment (Eastern Scholar) at Shanghai Institutions of Higher Learning (No. TP2014004), which are greatly acknowledged. We thank Anqi Hu from Fudan University for the assistance of AFM measurement.
The authors declare that they have no competing interests.
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Electronic supplementary material
The filter sets used for imaging four dyes inside HeLa cells; Lifetime measurement of the Eu clathrate in aqueous solution; Absorption and emission spectra of four dyes; Quantification of amino groups of BSA and cBSA; Emission spectra excited at 337 nm at different temperature; The emission spectra of the Eu clathrate at different pH; The influence of oxygen on the emission spectra of the Eu clathrate; The emission spectra of Rhodamine B at different pH; The influence of oxygen on the pH response of the triple nanosensor; The emission spectra of PtTCPP at different pH; Cell viability of HeLa cells co-cultured with nanosensors using CCK-8; Coomassie blue stained SDS-PAGE of BSA, cBSA and the triple nanosensor; The calibration plot of the nanosensor at different pH measured by fluorescent microscope.
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Zhang, X., Zhang, W., Wang, Q. et al. Quadruply-labeled serum albumin as a biodegradable nanosensor for simultaneous fluorescence imaging of intracellular pH values, oxygen and temperature. Microchim Acta 186, 584 (2019). https://doi.org/10.1007/s00604-019-3674-4
- Simultaneous imaging
- Multiple sensing
- Triple sensing
- Intracellular sensing
- Serum albumin
- Ratiometric pH sensing
- Lifetime imaging
- Oxygen sensing