Bioimaging Applications Of Human Serum Albumin Non-Covalently Bound Cyanine

Cyanine(Cy)dyes have gained widespread use in fluorescent biomolecular detection and disease diagnosis due to their near-infrared(NIR)absorption and emission properties.However,their susceptibility to oxidation and bleaching by reactive oxygen species,as well as easy aggregation and poor biocompatibility,limit their applicability in chemical biology and clinical laboratory diagnosis.One strategy to enhance the stability and biocompatibility of Cy dyes is through their non-covalent binding to Human Serum Albumin(HSA).This study presents a comprehensive investigation into the combination of two types of Cy dyes,Cy3 and Cy-Mu-7,with HSA,and the resulting improvement in photophysical properties and biological compatibility.Based on the idea that Cy can bind to HSA non-covalently,this paper conducted an in-depth study on the combination of two types of Cy dyes: Cy3(including P1,P2,P3 and P4)and Cy-Mu-7 with HSA.The HSA-Cy complex was successfully constructed by non-covalent binding of Cy with HSA,which achieved the photophysical properties and biological compatibility improvements of Cy properties,including fluorescence enhancement,enhanced fluorescence quantum yield,photostability improvement,lower aggregation tendency,higher solubility,and lower cytotoxicity.On this basis,the biological application of Cy dye was further expanded,including the diagnosis and treatment of tumors and the diagnosis of kidney diseases.The specific research work is as follows:Initially,four Cy3 dyes(P1,P2,P3,and P4)and one asymmetric cyanine dye,Cy-Mu-7,were synthesized and characterized.The UV-VIS absorption and fluorescence spectra of the system induced by the spontaneous combination of Cy3 and HSA in phosphate-buffered saline(PBS)were preliminarily studied.Spectroscopic analysis revealed that the combination of HSA and Cy3 significantly enhanced the UVvisible absorption signal and fluorescence emission signal of the system,providing a basis for the subsequent preparation of HSA-Cy complexes for imaging applications in living systems.Building upon this initial work,HSA-Cy3 nanoparticles were prepared by heating the complex formed by the spontaneous combination of HSA and Cy3.It was observed that heating induced albumin shrinkage and tight packing of Cy3(HSA-Cy-T),greatly improving the photostability of Cy3 fluorescent dyes.The resulting nanoparticles showed excellent photothermal effect under 660 nm laser irradiation,with higher cell uptake rates and lower dark and phototoxicity compared to P2 dye alone.Through cell and in vivo studies,HSA-P2-T was found to effectively eliminate cancer cells and ablate tumors under laser irradiation by enriching in the mitochondria of cancer cells.Additionally,the asymmetric cyanine Cy-Mu-7,with a large Stokes shift,was obtained through structural modification of conventional Cy7 dye.The effect of HSA on the optical properties of Cy-Mu-7 was investigated,revealing a longer fluorescence lifetime due to reduced non-radiative transition.This enabled fluorescence lifetime imaging to distinguish the fluorescence signals of Cy-Mu-7 and Cy-Mu-7-HSA and evaluate their respective sources of fluorescence signals in vivo.Both Cy-Mu-7 and Cy-Mu-7-HSA achieved fluorescence labeling of living kidney and liver in mice after injection into the tail vein,with the renal labeling properties of Cy-Mu-7-HSA being utilized for in vivo noninvasive diagnosis of ureteral obstruction.Overall,this study demonstrates the potential of HSA-Cy complexes for improving the stability and biocompatibility of Cy dyes,expanding their biomedical applications,including the diagnosis and treatment of tumors and kidney diseases.The preparation of HSA-Cy nanoparticles,along with the modification of Cy dyes,provides a promising avenue for the development of optical functional materials for living image analysis and diagnosis and treatment.