Hyo Won Lee

Development of Imidazoline‐2‐Thiones Based Two‐Photon Fluorescence Probes for Imaging Hypochlorite Generation in a Co‐Culture System

We designed and prepared the imidazoline-2-thione containing OCl(-) probes, PIS and NIS, which operate through specific reactions with OCl(-) that yield corresponding fluorescent imidazolium ions. Importantly, we demonstrated that PIS can be employed to image OCl(-) generation in macrophages in a co-culture system. We have also employed two-photon microscopy and PIS to image OCl(-) in live cells and tissues, indicating that this probe could have wide biological applications.

Ratiometric Two-Photon Fluorescent Probe for Quantitative Detection of β-Galactosidase Activity in Senescent Cells

We reported a ratiometric two-photon fluorescent probe (SG1) for β-galactosidase (β-gal) and its application to quantitative detection of β-gal activity during cellular senescence in live cells and in aged tissues. This probe is characterized by a significant two-photon excited fluorescence, a marked blue-to-yellow emission color change in response to β-gal, easy loading, insensitivity to pH and reactive oxygen species (ROS), high photostability, and low cytotoxicity. In addition, we show that SG1 labeling is an effective tool for quantitative detection of senescence-associated β-gal activity at the subcellular level in situ. This finding demonstrates that SG1 will find useful applications in biomedical research, including studies of cell aging.

Red emissive two-photon probe for real-time imaging of mitochondria trafficking.

Mitochondria trafficking plays an essential role for supplying energy in the neuronal system. We report here a red emissive two-photon probe for mitochondria (CMT-red) that showed high selectivity and robust staining ability for mitochondria, high photostability under a two-photon microscopy imaging condition, and low cytotoxicity. This probe can be easily loaded into live cells and tissue and used for real-time, high resolution imaging of the mitochondria trafficking in primary cortical neurons as well as in rat hippocampal tissue.

A ratiometric two-photon probe for Ca2+ in live tissues and its application to spinal cord injury model

Ratiometric imaging with a small-molecule probe is important for the in-situ quantitative analysis of chemical events. We developed a ratiometric two-photon fluorescent probe (SCa1-IREF) derived from dual dyes with different Stokes shifts. This probe has two identical windows: a Ca2+-sensing window and an internal reference window, with eliminated FRET interference. SCa1-IREF shows a marked change in the ratio upon response with Ca2+, significant two-photon brightness, considerable selectivity for Ca2+, and cell loading ability with low cytotoxicity. The ratiometric two-photon microscopy images revealed that this probe could directly and quantitatively estimate Ca2+ in live neurons and various tissues including rat spinal cord tissue. The studies of spinal cord injury model revealed that the Ca2+ level was significantly affected by elapsed time after injury. These results will provide useful applications for in-situ [Ca2+]i imaging and for the development of effective ratiometric probes.