Development and application of an integrated robotic imaging platform for cell culture monitoring and optimization

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Preamble: Characterization of cells and the development of clonal lineage heterogeneity through differentiation and epigenetic drift poses many fundamental challenges at the heart of bioprocess and tissue engineering. There is an urgent and specific need for the development of nondestructive multi-parameter analytical technologies that can assess cellular dynamics along trajectories that span multiple generations.      Objectives: The long term objective of my laboratory is to develop bioengineering strategies that enables the optimization and characterization of bioprocesses and products at the level of the individual cell.  Bioengineering at this level of detail requires robotic microscopy, single cell handling and manipulation, and an informatics approach to data analysis.  We hypothesized that cellular behavior analysis performed using long-term high resolution live-cell imaging will provide sufficient measures to enable culture control schemes that direct the development of individual cells along specific differentiation trajectories. Classification and control schemes will be limited only by the spatial-temporal-spectral resolution of imaging and the robustness of dynamic behavior and cell appearance identification.    Approach:  The work performed in this project will involve continued development and optimization of mechanical and photonic manipulation of individual cells for bio-assembly;  the implementation of multidimensional bright field and fluorescence time course imaging using high precision microscope robotics; the formulation of algorithms and software to analyze the multivariate data produced during extended time course imaging.  Each of these components will now be integrated into a robotic-imaging bioreactor system to enable the tracking and analysis of hundreds to thousands of cells at resolutions in which details are observable for each cell over time.Novelty:  The integration of high-content multidimensional live-cell imaging with informatics and single cell manipulation robotics and photonics represents a highly novel platform for biological discovery and the development of advanced bioprocess engineering methodologies.  Further, the application of lineage informatics for the analysis of cells represents an entirely new level of detail in culture analysis - cultures are not analyzed in terms of gross parameter measurements, but rather parameters are estimated and tracked at the single cell scale over multiple generations.  This work anticipates manufacturing challenges that will arise as cells are used for transplant therapy and single-cell analysis becomes a requirement.  As such, the proposed research program has the potential to develop fundamental methods and strategies that will find wide application in the areas of cell therapy and cell-based pharmaceutical manufacturing.