Development of a ratiometric time-resolved luminescence sensor for pH based on lanthanide complexes

Abstract

Time-resolved luminescence bioassay technique using lanthanide complexes as luminescent probes/sensors has shown great utilities in clinical diagnostics and biotechnology discoveries. In this work, a novel terpyridine polyacid derivative that can form highly stable complexes with lanthanide ions in aqueous media, (4′-hydroxy-2,2′:6′,2′′-terpyridine-6,6′′-diyl) bis(methylenenitrilo) tetrakis(acetic acid) (HTTA), was designed and synthesized for developing time-resolved luminescence pH sensors based on its Eu3+ and Tb3+ complexes. The luminescence characterization results reveal that the luminescence intensity of HTTA–Eu3+ is strongly dependent on the pH values in weakly acidic to neutral media (pKa=5.8, pH 4.8–7.5), while that of HTTA–Tb3+ is pH-independent. This unique luminescence response allows the mixture of HTTA–Eu3+ and HTTA–Tb3+ (the HTTA–Eu3+/Tb3+ mixture) to be used as a ratiometric luminescence sensor for the time-resolved luminescence detection of pH with the intensity ratio of its Tb3+ emission at 540nm to its Eu3+ emission at 610nm, I540nm/I610nm, as a signal. Moreover, the UV absorption spectrum changes of the HTTA–Eu3+/Tb3+ mixture at different pHs (pH 4.0–7.0) also display a ratiometric response to the pH changes with the ratio of absorbance at 290nm to that at 325nm, A290nm/A325nm, as a signal. This feature enables the HTTA–Eu3+/Tb3+ mixture to have an additional function for the pH detection with the absorption spectrometry technique. For loading the complexes into the living cells, the acetoxymethyl ester of HTTA was synthesized and used for loading HTTA–Eu3+ and HTTA–Tb3+ into the cultured HeLa cells. The luminescence imaging results demonstrated the practical utility of the new sensor for the time-resolved luminescence cell imaging application.