Fourier-filtering based size-encoded images for label-free tracking of sub-cellular organelles in neurons

Mitochondrial dynamics such as fission, fusion and movement can reveal the physiological status of neurons and, most importantly, serve as diagnostic measures for several neurodegenerative diseases. Traditionally fluorescent probes have been used to track mitochondrial dynamics in neurons. However, neurons show low transfection efficiency, presenting challenges for the use of genetically-encoded fluorescent markers. Alternatively, synthetic fluorescent dyes are shown to hinder mitochondrial motility. In addition, all types of fluorescent probes are subject to photo-bleaching which precludes imaging for longer periods of time. To circumvent these issues, we propose a light-scattering based label-free technique called Optical Scatter Imaging (OSI) that is sensitive to changes in morphology. In this work, we employ a previously reported label-free parameter to probe the change in the size of organelles such as mitochondria as they undergo fusion or fission in neurons. In addition, we present a technique to track organelle motion using kymographs obtained from the label-free images and compare them with those obtained from fluorescent images. We demonstrate that the label-free kymograph can track organelles such as mitochondria even after the sample is photo-bleached.