Preparation Of A New Type Of Near-infrared Fluorescent Probe And Its Application In Vivo

Near infrared fluorescence imaging is a novel,low-cost,controllable,high-sensitivity,high-throughput imaging method.It has advantages in detecting molecules,cells,animals and other aspects,and can make real-time observation of tumor cells and tissues in vivo in a non-invasive manner.In this paper,we plan to introduce functionalized structures based on the near-infrared fluorescent dye CyB-NO₃to prepare a near-infrared fluorescent probe with excellent optical performance and high biosafety,and further modify it with water solubility,so as to meet the application requirements of scientific research field and clinical stage.In this study,the near-infrared fluorescent dye CyB-NO₃ was prepared and synthesized,and maleimide structure was introduced to functionalize and construct a novel near-infrared fluorescent probe CyP with targeting ability.The near-infrared fluorescent probe was characterized by liquid chromatography,mass spectrometry,spectroscopy,solubility and other evaluation methods.The near infrared fluorescent dye CyB-NO₃ and the near infrared fluorescent probe CyP were modified by polyethylene glycol and bovine serum albumin(BSA),respectively,and the water-solubility and fluorescence properties were verified by gel electrophoresis.The cytotoxicity of CyB-NO₃ and CyP to tumor and normal cells,as well as the intracellular sublocalization and tracer effect were investigated at the cellular level.Finally,the acute toxicity of CyB-NO₃ and CyP in vivo and the fluorescence distribution in the whole body and tissues of mice were investigated at the small animal level,the excretion and tissue retention ability in vivo were analyzed,and the mouse model of lung cancer tumor transplantation was established to investigate the tumor accumulation effect of CyP in vivo.1.The preparation and characterization results of the fluorescent probe in this study show that the purity of CyB-NO₃ is?94%,the purity of CyP is?98%,the maximum ultraviolet absorption(?max)of CyB-NO₃ and CyP is 695 nm and 710/764nm,and the fluorescence emission wavelength(?em)is 788 nm and 794 nm,respectively.CyB-NO₃ and CyP have good fluorescence imaging effect and strong penetrating ability.In addition,the?max values of CyB-NO₃-PEG and CyP-BSA after water solubility modification of PEG and BSA based on CyB-NO₃ and CyP are 715/760nm and 700 nm,and?em are 790 nm and 797 nm respectively.2.The results of biosafety assessment showed that CyB-NO₃ concentration was greater than 10?m,and cytotoxicity was greater;The cytotoxicity of CyP in serum-containing system was low,and the biosecurity of CyP-BSA was high.The body weight of mice was not affected by 100?g of CyB-NO₃ and CyP,and the tissue sections of all organs were normal.3.The evaluation of imaging and tracer effect showed that CyB-NO₃,CyP and CyP-BSA all had high intensity fluorescence signals in the cytoplasm,and CyB-NO₃,CyP and CyP-BSA had high intensity fluorescence signals in mice with long duration,mainly distributed in liver and kidney and other organs.In addition,a mouse model of lung cancer tumor was established to further evaluate the imaging effect of CyP.It was found that CyP had a high degree of accumulation in the tumor sites of the tumor mice,and there were high intensity fluorescence signals.In summary,the near-infrared fluorescent dye CyB-NO₃ prepared in this study has excellent fluorescence characteristics,water soluble modification and slightly low biosafety.The near-infrared fluorescent probe CyP has the advantages of excellent fluorescence characteristics,safety,good imaging effect and high degree of aggregation of tumor tissue,which can be developed as a targeted probe for tumor diagnosis in vivo.