The Latest on Live Cell Imaging

A Head-to-Head feature with Mount Sinai Hospital and Nanopoint

“Providing tight control of the cell environment is critical in successful live cell imaging experiments”
-Cathy Owen, Nanopoint

NGP. Over the past decades, advances in live cell imaging have dramatically transformed the biological sciences. What have been the most critical of these advances, and how have they affected the field?
Laurence Pelletier. By far the most important discovery was the work in the laboratories of Shimomura, Chalfie, Inoue, Tsien and many others that led to the structure, expression, and finally optimization of the autofluorescent proteins. Live cell microscopy would have remained in the hands of a small number of skilled cell biologists had not these groups devised a simple way to use molecular biology to tag vital proteins in living cells. Gene products can be localized not only structurally, as we had with antibody technology, but also dynamically within living cells. With this technology, it is now possible to place virtually any biomolecule in the context of structure and dynamics; that is, within space and time.

Cathy Owen. Providing tight control of the cell environment is one of the most critical factors in successful live cell imaging experiments. Robust on-stage environmental controls along with automated perfusion of media ensure the viability of live cells when investigations require extended time lapse imaging sequences, and when serial addition of a drug and/or reagent must be accomplished without disrupting the cells. The technique of imaging live cells on the microscope stage is improved with the emergence of synthetic fluorophores and fluorescent proteins to serve as qualitative and quantitative reporters of intracellular structure and dynamics. In key areas of drug discovery, such as screening of compounds and preclinical testing of drugs in living cells, microfluidic systems make a useful contribution, and represent an improvement on existing technologies. Novel reaction, manipulation and analytical steps can be performed with microfluidic systems that are not accessible to other approaches providing new information about biological systems.

NGP. What special challenges are involved in live cell imaging, as distinct from other types of imaging?
CO. It can be argued that optical microscopes, cheap and readily-available computing power, sophisticated image analysis software, megapixel digital camera systems and a plethora of highly specific fluorescent tags mean that live cell studies today are rather straightforward and easy. There are still some technical challenges however. One of the most significant challenges for performing successful live cell imaging experiments is to maintain the cells in a healthy state and functioning normally on the microscope stage for an extended period of time while being illuminated in the presence of the fluorescent tags. Some live cell experiments, such as documenting the slow growth and reproduction of cells, may last for many days. In this case, just keeping the cells alive is a challenge that many live cell imaging workstations cannot meet and maintaining cells in an optimum physiological condition throughout the long observation phase is critical to the success of those experiments.

LP. In most applications, imaging is limited by dynamic range, detection and signal-to-background. In most cases these can be overcome by increasing exposure, making the signal brighter by increasing the illumination on the sample, or by changing the chemistry around the targets of interest. In live cell imaging, the biology has to drive the experiment. The timing of events and cell viability limit the exposure that can be used. Cells are stressed by exposure to light and the physiology within the cell will only tolerate so much manipulation.

The intent is generally to understand cell biology, not the biology of a dying cell. It is not enough to image the cell into apoptosis. Another challenge is the storage and the analysis of complex phenotypes using automated means. Again we will see different fields join forces to get at the problem (physics, optics, cell biology and informatics)

NPG. Why is it important that researchers from different disciplines interact to ensure progress in the field?
CO. If we are ever to achieve the goal of personalized medicine, it’s critical to move to a more “systems-biology” approach to research. Understanding the complexity of human disease and providing novel therapeutics requires the commitment of an interdisciplinary team of biologists, engineers, physicists, chemists, mathematicians, physicians, and computer scientists to tackle the difficult technical challenges. In addition to our interdisciplinary team, Nanopoint's extended time lapse live cell imaging solutions have been developed utilizing collaborations with reagent companies, biomedical researchers, and microfluidics experts. These types of collaborative partnerships enable new types of cellular experimentation that are essential to understanding complex human diseases, accelerating developments in stem cell research and pharmacogenomics, and breakthroughs in new treatments.

LP. We see more and more labs from a variety of disciplines making use of live cell imaging. Fifteen years ago, the field was limited to a handful of skilled microscopists and bioengineers. Now, virtually every field of biology uses live cell imaging. It is only through collaboration that scientists can adapt methods from all disciplines to optimize these experiments and to push the boundaries. The latest work in imaging borrows heavily from astrophysics, signal theory, spectroscopy and even quantum physics.

NGP. Working with and imaging live cells can be a complex task for microscopists who are unfamiliar with the process. What tools and techniques are available to help?
LP. Good instrumentation and mentorship! The systems that consistently yield superior results are built on a lot of knowledge, research and experience. Good companies are constantly making improvements to every aspect of their imaging systems to optimize them for imaging living cells. Secondly, I would highly recommend one of a number of excellent courses available for learning these techniques. Some of the top microscopists in the world regularly take the time to teach at these courses and there is no better way to quickly assimilate all that is needed to do these experiments well.

CO. Nanopoint has developed a tightly integrated system that provides new standards of precision and levels of efficiency for the study of individual and small groups of live cells, enables new approaches for multiple cell analysis and simultaneous processing, and supports multi-day time lapse imaging of live cells. The system integrates lab-on-a-chip technology fabricated from glass which increases the number and types of applications the cellTRAY® can be used for. Glass offers the low thermal conductivity and optical clarity that is ideal for live cell imaging applications. The cellTRAY Manager software provides the navigation, camera, shutter and filter controls, auto-focus, and microfluidics control necessary for today’s demanding live cell imaging applications. The user can automatically acquire images and save those images for further analysis by third party products such as ImageJ, MetaMorph, ImagePro, CellProfiler and others.

At the Samuel Lunenfeld Institute, Mount Sinai Hospital, Toronto, Laurence Pelletier’s lab studies cell cycle regulation of centrosome function and cilia formation in specialized cells. Functional genomics and cutting-edge live-cell microscopy are used to identify and study novel proteins required for these processes and elucidate their role in development and diseases

Cathy Owen is CEO, President and Director of Nanopoint. She brings more than 30 years of executive leadership, general business and entrepreneurial experience to Nanopoint. She joined the company in 2004 after a successful 21-year career at IBM, executive management of a Silicon Valley startup and president of a Hawaii software startup company.