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In recent years, organic fluorophores-based molecular probes with dual-fluorescence ratiometric responses to in-vitro / in-vivo pH (DFR-MPs-pH) have been attracting much interest in fundamental application research fields. More and more scientific publications have reported the exploration of various DFR-MPs-pH systems that have unique dual-fluorescence ratiometry as the signal output, in-built and signal self-calibration functions to improve precise detection of targets. DFR-MPs-pH systems possess high-performance applications in biosensing, bioimaging and biomedicine fields. This review has comprehensively summarized recent advances of DFR-MPs-pH for the first time. First of all, the compositions and types of DFR-MPs-pH are introduced by summarizing different organic fluorophores-based molecule systems. Then, construction strategies are analyzed based on specific components, structures, properties and functions of DFR-MPs-pH. Afterward, biosensing and bioimaging applications are discussed in detail, primarily referring to pH sensing and imaging detection at the levels of living cells and small animals. Finally, biomedicine applications are fully summarized, majorly involving bio-toxicity evaluation, bio-distribution, biomedical diagnosis and therapeutics. Meanwhile, the current status, challenges and perspectives are rationally commented after detailed discussions of representative and state-of-the-art studies. Overall, this present review is comprehensive, in-time and in-depth, and can facilitate the following further exploration of new and versatile DFR-MPs-pH systems toward rational design, facile preparation, superior properties, adjustable functions and highly efficient applications in promising fields. Schematic illustration of this present review about organic fluorophores-based molecular probes with dual-fluorescence ratiometric responses on in-vitro/in-vivo pH for biosensing, bioimaging and biotherapeutics applications. [Display omitted] • This review summarizes organic fluorophores-based molecular probes with ratiometric fluorescence biosensing of pH. • Compositions, constructions, biosensing, bioimaging and biomedicine applications of the probes are summarized. • Current status, challenges and perspectives are discussed. [ABSTRACT FROM AUTHOR]

Organic small molecule fluorescent groups have injected new material support into the field of medical imaging due to their unique luminescence mechanism and easy tuning of structure. The great potential of NIR-II window imaging forces us to continuously optimize the structure of organic fluorophores to design better fluorescent molecules for fluorescence imaging-guided surgery. An ideal organic small molecule fluorescent group: it can penetrate into the inside of the organism, clearly present the internal structure and the edge contour of different tissues, so as to perfectly achieve internal imaging and accurately guide external surgery. In vivo, fluorescent groups do not damage normal tissues and organs. However, problems such as low quantum yield and poor biocompatibility greatly limit the clinical transformation of NIR-II fluorescent small molecules. To avoid the shortcomings of NIR-II fluorescent probes as much as possible and better realize image-guided surgery, in this experiment, the biplane donor unit was incorporated into the twisted D-π-A-π-D structure to expand the conjugated structure of the fluorescent group, which not only realized NIR-II emission, but also had high quantum yield and biosafety. [Display omitted] • CED NPs have high quantum yield and biosafety. • CED NPs can clearly show the tumor contour of subcutaneous tumor nude mice. • CED NPs can clearly label the enlarged lymph nodes in nude mice with metastatic tumors. [ABSTRACT FROM AUTHOR]