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3D Orbital Tracking in a Modified Two-photon Microscope: An Application to the Tracking of Intracellular Vesicles

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How to track protists in three dimensions.

Knut Drescher¹, Kyriacos C Leptos, Raymond E Goldstein

¹Department of Applied Mathematics and Theoretical Physics, Centre for Mathematical Sciences, University of Cambridge, Wilberforce Road, Cambridge CB3 0WA, United Kingdom.

The Review of Scientific Instruments

|February 5, 2009

Summary

This summary is machine-generated.

We developed a 3D tracking system for microorganisms (10-1000 microm) with minimal fluid motion. This apparatus enables detailed study of phototaxis and surface interactions for various species.

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Area of Science:

Biophysics
Microfluidics
Microorganism behavior

Background:

Accurate tracking of microorganisms is crucial for understanding their behavior.
Previous methods faced limitations in 3D tracking, proximity to surfaces, and fluid motion interference.

Purpose of the Study:

To present a novel apparatus for high-resolution 3D tracking of swimming microorganisms.
To enable the study of microorganism behavior without surface or fluid motion artifacts.
To facilitate detailed analysis of phototaxis and other taxes.

Main Methods:

Utilizing two synchronized, perpendicularly oriented microscopes with CCD cameras.
Employing narrow-band illumination to prevent phototactic responses.
Combining images for precise 3D trajectory reconstruction.
Using directional illumination with millisecond timing for dynamic studies.

Main Results:

Achieved 3D tracking of microorganisms in the 10-1000 micrometer size range.
Minimized background convective fluid motion for accurate measurements.
Successfully studied phototactic trajectories of various microorganisms, including Chlamydomonas and Volvox.
Enabled investigation of surface-mediated hydrodynamic interactions.

Conclusions:

The developed apparatus provides a robust platform for studying microorganism motility and behavior in controlled environments.
The system allows for detailed analysis of light-mediated responses and hydrodynamic interactions.
Minimal modifications permit future studies on chemotaxis and other stimulus-response behaviors.