Rational Engineering a Multichannel Upconversion Sensor for Multiplex Detection of Matrix Metalloproteinase Activities

Abstract

Optical sensing of cancer-relevant protease is of great value for cancer diagnostics, prognosis, and drug discovery. Multiplex sensing is known to improve predicative accuracy yet remains challenging because of severe fluorescence signal crosstalk in a single assay. Herein, we developed a multichannel optical sensor based on upconversion nanoparticles for multiplex ratiometric sensing of proteolytic activities of two matrix metalloproteinases (MMP-2 and MMP-7). To this end, we rationally designed a NaYF₄:Gd³⁺/Yb³⁺@NaYF₄:Yb³⁺/Tm³⁺/Er³⁺ core-shell structure that favors multicolor narrow-band emission of both Tm³⁺ and Er³⁺ dopants and efficient luminescence resonance energy transfer (LRET) between the dopants in the shell and the fluorophores on the particle surface. The sensor was constructed via a facile phase transfer protocol using two polyhistidine-containing peptides conjugated with different fluorophores (FITC and TAMRA) as coligands. The blue and green emission could be specifically activated by MMP-7 and MMP-2, respectively, upon peptide cleavage, and the red emission could serve as an internal reference for ratiometric sensing. The sensor exhibits high specificity and sensitivity toward both targets with little signal crosstalk and cross-reactivity. It could potentially serve as a general platform for multiplex detection of various types of proteases.