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Improved Neuronal Performance with the Gibco B-27 Plus Culture System

Speaker
  • Director and Group Leader, Thermo Fisher Scientific
    Biography
      Dr. Kuninger leads research, development and commercialization of media systems for pluripotent stem cell culture & differentiation, neurobiology, and (non-hepatic) primary cell biology at Thermo Fisher Scientific in the Cell Biology business based in Frederick MD. His teams support numerous portfolios and have launched over 25 new products spanning stem cell culture & cryopreservation, differentiation (endo-, ecto- and meso-dermal lineages) and neurobiology over the past 3 years. David is a seasoned scientist and manager, experienced in media formulation & optimization, assay design and implementation, and troubleshooting. Expertise in GLP/GMP compliance, tech transfer and scale up, as well as verification and validation processes. Prior to starting at Thermo Fisher Scientific (legacy Invitrogen) in 2007 as Staff Scientist, he joined Oregon Health Sciences University as a Postdoctoral Fellow investigating the actions of insulin-like growth factors in the lab of Dr. Peter Rotwein, subsequently joining the faculty in the Department of Biochemistry at OHSU as a Research Instructor. He completed is PhD in Biochemistry and Genetics University of Texas Medical Branch in the laboratory of Dr. John Papaconstatinou and has a B.S. in Chemistry from the University of Oregon.

    Abstract

    Human iPSC-derived neurons have increasingly become a valuable system for the study of neurological disorders.   Robust cell reprogramming and improved differentiation protocols enable scientists to generate patient-specific, disease in a dish models for disorders such as Parkinson's, Alzheimer's, and Autism, among others.  These human models tend to be flexible, scalable and maintain many of the characteristics of found in these disorders, which are key requirements for their use in mechanistic and drug discovery studies.  Further, the development of gene editing technology has spawned intense interest in the use of gene-edited, patient-specific iPSC-derived neurons in cell therapy applications for the treatment of neuro-degenerative disorders.

    A critical step in generating useful iPSC-derived neurons is neuronal maturation.  During maturation neurons extend neurites to form highly connected networks, express synaptic markers, and become electrically active.  Typical maturation conditions are inefficient,  generating poorly matured neurons with low levels of functionality over extended periods of time.  Recently we developed a new neuronal neuronal maturation and maintenance system, (B-27TM Plus and NeurobasalTM Plus) and showed significantly improved neuronal survival, maturation, and functionality of primary rodent neurons compared to other culture systems.

    Here we expand our studies to PSC-derived neurons, utilizing multiple human lines PSC and different approaches for neural stem cell derivation.  Diverse endpoints were used to interrogate maturation; neurite outgrowth, neuronal maturation marker expression (through quantitative imaging), and functionality through Multi-Electrode Array (MEA) analysis.  We found that human PSC-derived neurons matured in the new “Plus” system showed both accelerated neurite outgrowth and improved activity as compared to other approaches.   Additionally our studies highlight the importance of optimizing several key parameters, including extracellular matrix coating concentrations and delivery conditions for improved reproducibility and quality of stem cell derived neural cultures.


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