Snapshot 3D tracking of insulin granules in live cells

TL;DR

This paper introduces a multifocal microscope technique for rapid 3D imaging and tracking of insulin granules in live cells, revealing insights into their motion and cytoskeletal interactions.

Contribution

The study presents a novel 3D snapshot imaging method using MFM for accurate tracking of insulin granules in live cells, enabling detailed analysis of their dynamics.

Findings

Granules exhibit super-diffusive motion in 3D

Motion is less super-diffusive in 3D than 2D, indicating cytoskeletal anisotropy

Validated imaging accuracy with fluorescence beads

Abstract

Rapid and accurate volumetric imaging remains a challenge, yet has the potential to enhance understanding of cell function. We developed and used a multifocal microscope (MFM) for 3D snapshot imaging to allow 3D tracking of insulin granules labeled with mCherry in MIN6 cells. MFM employs a special diffractive optical element (DOE) to simultaneously image multiple focal planes. This simultaneous acquisition of information determines the 3D location of single objects at a speed only limited by the frame rate of array detector . We validated the accuracy of MFM imaging and tracking with fluorescence beads; the 3D positions and trajectories of single fluorescence beads can be determined accurately over a wide range of spatial and temporal scales. The 3D positions and trajectories of single insulin granules in a 3.2 micro meter deep volume were determined with imaging processing that combines 3D decovolution, shift correction, and finally tracking using the Imaris software package. We find that the motion of the granules is super-diffusive, but less so in 3D than 2D for cells grown on coverslip surfaces, suggesting an anisotropy in the cytoskeleton (e.g. microtubules and action).