In vivo tracking and measurement of pollen tube vesicle motion

Particle tracking has emerged as a powerful tool for investigating the swarm control of microrobots and the dynamic biological processes in the life sciences. In seed plants, pollen tubes, a part of the male gametophyte, are excellent models for understanding plant growth and cellular behavior, because vesicle motion within pollen tubes reveals important information about vesicle function and interactions. Conventional vesicle tracking is based on spatiotemporal image analysis, which requires high-quality images and vesicles with constant velocity. For in vivo tracking, vesicles may disappear in some frames, and image sequences may have spatial and temporal distortions, which hamper vesicle tracking for broader applications. In this paper, we studied intracellular motion during pollen tube growth with an optical flow method. Streaming images from confocal and optical microscopes were recorded to study the intracellular motion of vesicles of different size. Local motion for each vesicle was detected using a local displacement vector field. The displacement from two adjacent frames was then calculated. The flow field shows information such as the dynamics of vesicle secretion, endocytosis, exocytosis, and cytoskeletal stability. Vesicles from different regions inside the tube were tracked simultaneously with a Kanade-Lucas-Tomasi (KLT) feature matching algorithm. The spatial and temporal characteristics of intracellular vesicles were evaluated. The proposed methods can be of great use for studying the dynamics of fluorescently tagged particles in biological systems.