Lighting up ATP in cells and tissues using a simple aptamer-based fluorescent probe

Xuena Zhu, Nanobioengineering/Bioelectronics Laboratory, Department of Biomedical Engineering, Florida International University; Department of Pathology, the First Affiliated Hospital and School of Medicine, Zhejiang University
Wenjun Liu, Zhejiang Provincial Key Laboratory of Pancreatic Disease and Department of Hepatobiliary and Pancreatic Surgery, the First Affiliated Hospital, Zhejiang University School of Medicine
Maedeh Mozneb, Nanobioengineering/Bioelectronics Laboratory, Department of Biomedical Engineering, Florida International University
Chen-Zhong Li, Nanobioengineering/Bioelectronics Laboratory, Department of Biomedical Engineering, Florida International University; Center for Cellular and Molecular Diagnostics, Department of Biochemistry and Molecular Biology, Tulane University School of Medicine

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Abstract

Extracellular ATP as a purinergic signaling molecule, together with ATP receptor, are playing an important role in tumor growth, therapy resistance, and host immunity suppression. Meanwhile ATP is a crucial indicator for cellular energy status and viability, thus a vital variable for tissue regeneration and in vitro tissue engineering. Most recent studies on COVID-19 virus suggest infection caused ATP deficit and release as a major characterization at the early stage of the disease and major causes for disease complications. Thus, imaging ATP molecule in both cellular and extracellular contexts has many applications in biology, engineering, and clinics. A sensitive and selective fluorescence signal-on probe for ATP detection was constructed, based on the base recognition between a black hole quencher (BHQ)-labeled aptamer oligonucleotide and a fluorophore (Cy5)-labeled reporter flare. The probe was able to detect ATP in solution with single digit mu M detection limit. With the assistance of lipofectamine, this probe efficiently entered and shined in the model cells U2OS within 3 h. Further application of the probe in specific scenery, cardio-tissue engineering, was also tested where the ATP aptamer complex was able to sense cellular ATP status in a semi-quantitative manner, representing a novel approach for selection of functional cardiomyocytes for tissue engineering. At last a slight change in probe configuration in which a flexible intermolecular A14 linker was introduced granted regeneration capability. These data support the application of this probe in multiple circumstances where ATP measurement or imaging is on demand.