Bio-functional Dye Derived Fluorescent Probe And Live Cell Imaging Based Research

Based on benzothiazole, naphthalimide and heptamethine cyanine dyes, designed and synthesized three sensitive fluorescence probes which applied mechanisms include: Excited-State Intramolecular Proton Transfer(ESIPT), fluorescence polarization technology(FP) and dual excitation, dual emission near-infrared fluorescence response. Living-cell imaging research was also conducted.(1) Designed and synthesized a first ratiometric fluorescent probe PDS-2 for ratiometric detection of changes of periodate based on ESIPT mechanism. This PDS-2 based fluorescent technique may enable for a better understanding of periodate related biological and chemical processes. Also, it is an efficient tool for public health, food safety and environmental protection.(2) A first naphthalimide based fluorescence polarization probe BIO was designed and synthesized for quantitive detection of cancer cells. Fluorescence polarization technology has self-calibration function which enable detection more accurate. The detaction process is in homogeneous solution without cell lysis and further separation steps. Experiments showed that the lowest detection limit of BIO for He La cells was about 85 cells m L-1, the linear range was from 2.5 × 102 cells m L-1 to 1 × 106 cells m L-1. Naphthalimide has relatively longer excited-state lifetime and higher photostability that makes BIO particularly suitable for living-cell imaging under continuous irradiation with confocal microscopy, and the specific interaction of BIO with CD44-overexpressing cell lines was clearly visualized by living-cell imaging.(3) Based on heptamethine cyanine dye owns advantages of long wavelength of near-infrared emission, rigid plane structure which enable probe to have several binding site and relative high affinity constant to proteins, we also combine that serum albumins have structure-selective hydrophobic cave whose small polar environment leads enhancement of fluorescence intensity to design and synthesize one cyanine dye fluorescent probe C7 H, through its dual wavelengths turn-on of emission signals after binding with HSA, experiment results indicated that synthesized target probe has relative high site-specific affinity to serum albumins. It could realize quantitative analysis for HSA, and also applied it in human urine and serum for HSA detection of real samples.(4) We applied synthesized FRET ratiometric probe VTAF to carry on research on visible dynamic tracing of vicinal dithiol-containing proteins(VDPs) in living cells. Experiment results suggested that there are abundant VDPs in mitochondria. Moreover, some redox-sensitive VDPs are also present on cell surface which can respond to redox stimulus. This ratiometric fluorescence technique could realize trapping and quantifying protein vicinal dithiols in living cells, as well as its visible dynamic tracing of VDPs.