Fluorescence Lifetime Imaging of p-tau Protein in Single Neuron with a Highly Selective Fluorescent Probe

Hyperphosphorylated tau (p-tau) protein is one of the key markers of Alzheimer's disease (AD). However, a lack of analytical methods for p-tau protein with high selectivity and accuracy is the bottleneck to understand the pathological processes of AD. In the present work, a highly selective fluorescence lifetime imaging microscopic (FLIM) probe (τ-p-tau) was rationally designed and developed for determination of p-tau protein, in which binuclear zinc units were designed as the recognition groups, and an improved cyanine was synthesized as the dye unit. The FLIM measurement is independent of the concentration of fluorescent probes, irradiation light sources, and measurement environments. Meanwhile, the developed τ-p-tau probe demonstrated strong affinity toward p-tau protein, thus enhancing the selectivity of determination of p-tau protein against Aβ, Aβ fibril, tau, protein kinases, ATP, and so on. The fluorescence lifetime of τ-p-tau probe showed a good linearity with the concentration of p-tau protein from 1.0 to 5.0 μM with a low detection limit of 85.15 ± 0.03 nM. The response time of the present probe for p-tau protein was estimated to be less than 6.2 s. Furthermore, taking the advantages of low toxicity and good biocompatibility, the developed probe was successfully applied to monitor the fluctuation of concentration of p-tau protein by FLIM imaging at single neuron level. It was found that the concentration of p-tau protein inside live neurons obviously changed upon oxidative stress.