Engineering a perfusion-enabled mechanical compressor for long-duration immobilization and microscopy of cells and small organisms
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Date
2011-04
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Vanderbilt University. Dept. of Physics and Astronomy
Abstract
The study of living specimens is essential to the understanding of
organismal behavior. Unfortunately, a major difficulty in the study of
live organisms is that many move in and out of the field of view or
focal plane during microscopy. The present work seeks to combat this
considerable problem by developing a mechanical microcompressor that
immobilizes living cells and small organisms for long-duration optical
microscopy. The device, dubbed the "Commodore Compressor," features
two key innovations: (1) the integration of a perfusion system to keep
the trapped specimen alive over several hours, as well as permitting
the addition of chemoattractants, drugs, and other chemicals; (2) the
incorporation of an optional patterned PDMS platform to improve the
efficacy of immobilization in a targeted, organism-specific manner.
One application of the Commodore Compressor is in monitoring the
change in protein bioluminescence intensity in many trapped
Saccharomyces cerevisiae cells during synchronized cell cycles. The
experiment's feasibility and key techniques have been well
demonstrated, although bioluminescence cannot currently be visualized.
A second application involves fluorescence imaging of the neural
network development of immobilized Caenorhabditis elegans over many
hours. The development of new patterned PDMS platform designs, aided
by the innovative use of established techniques, has driven the
present work toward accomplishing the goal, but true long-term
viability remains elusive. The Commodore Compressor may be directly
used or easily adapted for many other specimen types and experimental
scenarios.
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Keywords
compressor, immobilization, microscopy, microfabrication, polydimethylsiloxane, PDMS, Caenorhabditis elegans, Saccharomyces cerevisiae, yeast, nematode