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Comprehensive Characterization of Pluripotent Stem Cells

Speaker
  • Staff Scientist, Thermo Fisher Scientific
    Biography
      Dr. Pradhan joined Thermo Fisher scientific in 2016 and is currently working on next generation tools and technologies development with the goal of providing assays and methods for molecular characterization of stem cells and T cells utilizing methylation platform, array-based whole genome and transcriptomic profiling and TaqMan qPCR assays. He earned his Ph.D. in Molecular Biophysics from University of Calcutta, India, investigating the structural and functional consequences of binding of putative anti-cancer small molecules to chromatin in the context of transcription and disease. After graduating Dr. Pradhan moved to United States and did two postdocs -- a brief one at UC San Diego followed by a second postdoc at UC Los Angeles. He made a few seminal contributions in unraveling the mechanism of eukaryotic gene expression and silencing leveraging sophisticated biochemistry and extensive next generation sequencing approaches.

    Abstract

    As reprogramming methodologies have become more reliable and efficient, corresponding improvements were needed in the characterization workflow as well; specifically in terms of the speed, efficiency, specificity and sensitivity of the methods used.  Molecular analysis platforms offer a quantitative, accurate, and fast alternative to current methods and have recently been utilized to qualify pluripotent stem cells (PSC). When characterizing stem cells, it is important to confirm that a cell line a)  has not acquired any genomic abnormalities during derivation or genetic engineering, b) has or (maintains) the  capacity to differentiate into all three germ layers, and c) maintains expression of pluripotency associated markers. In addition to the characteristics described above, it is highly desirable to be able to uniquely ID every parental and subsequent derivative cell line; especially in core facilities, stem cell banks, or cell therapy development.
    Here we describe three characterization methods: PluriTest, KaryoStat, and ScoreCard,. PluriTest and KaryoStat leverage microarray based platforms that provide a comprehensive gene expression profile to determine the state of pluripotency and whole genome coverage to detect chromosomal abnormalities via copy number variations, respectively. ScoreCard utilizes a focused panel of TaqMan qPCR based assays to assess expression of specifically selected genes that confirm pluripotency and differentiation potential.  These three methods provide a complete solution to fully characterize PSC lines for pluripotency, differentiation potential, genomic integrity and sample ID. 


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