Ratiometric Fluorescent Probe for Imaging of Pantetheinase in Living Cells

Pantetheinase, which catalyzes the cleavage of pantetheine to pantothenic acid (vitamin B5) and cysteamine, is involved in the regulation of oxidative stress, pantothenate recycling and cell migration. However, further elucidating the cellular function of this Enzyme is largely limited by the lack of a suitable fluorescence imaging probe. By conjugating pantothenic acid with cresyl violet, herein we develop a new fluorescence probe CV-PA for the assay of pantetheinase. The probe not only possesses long analytical wavelengths but also displays linear ratiometric (I_{628/582 nm}) fluorescence response to pantetheinase in the range of 5-400 ng/mL with a detection limit of 4.7 ng/mL. This probe has been used to evaluate the efficiency of different inhibitors and quantitatively detect pantetheinase in serum samples, revealing that pantetheinase in fetal bovine serum and new born calf serum is much higher than that in normal human serum. Notably, with the probe the ratiometric imaging and in situ quantitative comparison of pantetheinase in different living cells (LO2 and HK-2) have been achieved for the first time. It is found that the level of pantetheinase in LO2 cells is much larger than that in HK-2 cells, as further validated by Western blot analysis. The proposed probe may be useful to better understand the specific function of pantetheinase in the pantetheinase-related pathophysiological processes.