Imaris Customer Profile

Cellular mechanics by Department of Biological Sciences, National University of Singapore (NUS)

Cellular mechanics has been a focus of Dr. Paul Matsudaira’s research for more than a decade. This leading scientist uses and develops powerful microscopy imaging techniques and complex analysis methods to study how cells can carry out complex tasks such as coordinated and three-dimensional movement.

Dr. Matsudaira is currently at the National University of Singapore, where he heads the Department of Biological Sciences, directs the centre for BioImaging Sciences, co-directs the Mechanobiology Institute, and is a professor in the Division of Bioengineering. He is also a co-author of the popular Molecular Cell Biology textbook published by W.H. Freeman. From 2001 to 2008, Dr. Matsudaira directed the Whitehead Institute/MIT Center for BioImaging, and it here that his lab began studying the mechanics of cell motility and developing methods to visualize this motility through imaging.

In an early study, Dr. Matsudaira and Ivan Correia examined the dynamics of cell adhesion.1 As a cell such as a macrophage moves forward, its leading edge protrudes and adheres to the substrate. They used Imaris to identify colocalized proteins, which let them pinpoint a specific interaction that took place between components of actin and intermediate filament cytoskeletons during the adhesion process.

This was followed by a study in which James Evans and Correia, who were postdocs in Dr. Matsudaira’s lab, studied the assembly of podosomes during macrophage movement.2 Podosomes are projections at the leading edge of a macrophage that contain actin and fimbrin.

The researchers extracted detailed kinetic information about podosome dynamics. They used Imaris neuron tracer to track podosomes as they appeared and disappeared, and then stitched together 2-D time-lapse images for a full picture. They used the data to form a kymograph that allowed visualization of dynamics occurring at the leading edge. “Kymography is a visualization method that dates from the early 1900s and is now a valuable tool in cell biophysics visualization,” says Dr. Matsudaira. This work introduced them to the mechanics of cell migration in two and three dimensions.


Mechanical Integrity
Biologists are very interested in understanding how cells handle the forces under which they operate in the body. For example, cytoplasmic gel must sustain shear stresses of up to 1,000 Pa for proper cell function. F-actin plays an important role in maintaining a cell’s mechanical integrity when undergoing various stresses, but it doesn’t act alone. In vivo actin filaments join into bundles or networks and work with actin binding proteins to influence the cells shape, division, adhesion, and motility.

Dr. Matsudaira was part of a team of researchers that examined how changes in the cell’s mechanical integrity correlate with cell structure.3 The researchers wanted to measure how the cell’s actin networks respond to increasing concentrations of the rigid actin-binding protein scruin. To directly visualize the actin network’s structure on various scales required electron microscopy, multiparticle tracking, and confocal microscopy.

Using Imaris, the researchers assembled confocal fluorescence microscopy images into 3-D projections that showed the bundled and cross-linked F-actin network. They calculated the actin network’s mesh size by measuring the peakto- peak distance in the intensity profile obtained across fluorescent images. The confocal information was correlated with that from the other techniques to provide quantitative information about how the structure of the actin network changes with varying scruin concentration.

3D Tissue
To truly understand cells and the multitude of processes in which they are involved requires studying them in a setting that closely replicates living tissues. Tissues have multiple types of cells, an extra-cellular matrix, and take in signals from other parts of the body. All these components work together to determine tissue function.

Matsudaira and his colleagues developed a way to create synthetic tissue that has the 3-D complexity of a multicellular organism.4 They assembled individual cells of varying types into a precise three-dimensional configuration by using multiple time-shared optical tweezers and microfluidics to organize a mixture of cell types into a complex network. Smaller microarrays of cells in a hydrogel microstructure are formed and then tiled together into a larger superarray. Using Imaris the researchers formed 3-D reconstructions from confocal images of the arrays. This let them examine the 3-D nature of the smaller microarrays as well as the superarrays.

“Imaris has always represented the state-of-art in visualization methods and has extended to include data measurement tools,” says Dr. Matsudaira. “We have worked with Bitplane over the past decade to introduce useful tools for biologists starting with the ability to tackle very large image datasets and batch processing methods. The visualization and quantitation tools have improved tremendously during this period, and we continue to employ Imaris in our research.”