Single-molecule tracking

Research Summary

In this work, we reported a single-molecule tracking approach to study the single-molecule dynamics of nuclear SOD1 in response to oxidative stress in neuron cells. We used H1-SOD1-mEos4b, as our model system, based on CRISPR-Cas9 technique. We have proven that the knock-in cell line was effective based on the sequencing and protein expression analysis of SOD1-mEos4b and we have observed that the engineered homozygous clone that produces SOD1-mEos4b fusion proteins displays typical stem cell morphology. The SOD1 localization and motion was captured by the fluorescent signals emitted from mEos4b and processed for further analysis. The trajectory and diffusion coefficient analysis showed that more SOD1 molecules were immobilized under H2O2 treatment compared to basal condition, which we proposed this could due to more SOD1 binding events happened during the oxidative stress response and therefore slower the nuclear SOD1 diffusion behavior.