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2020 Jun 16 - Knowledge Resource Series#2 Gene Therapy Webinar
Christopher Bunker, Ph.D.
Senior Director, Business Development, Advanced Cell Diagnostics
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Outline:

Visual detection and quantification of AAV and LV vector biodistribution and transgene expression in preclinical animal models with RNAscope® and BaseScope™ in situ hybridization. RNAscope in situ hybridization (ISH) has been described by its users as a means of “morphology-based AAV biodistribution analysis.” RNAscope in situ assays of AAV DNA and transgene mRNA combine the molecular sensitivity of qPCR with single-cell resolution and the context of tissue morphology. By combined RNAscope probes to AAV vector and transgene mRNA, vector copy number and expression can be visually assessed and quantified simultaneously in each cell of an intact fixed tissue. With the addition of cell marker probes in multiplex fluorescent RNAscope assays, AAV vector and transcript levels can be measured in any target cell type. BaseScope ISH is another powerful tool facilitating gene therapy development. Specific measurement of therapeutic human transgene expression in animal models is often impossible by IHC because of the high level of homology between transgene and animal ortholog transcripts. BaseScope enables specific detection of even single nucleotide differences and can easily differentiate AAV human transgene from non-human primate and other animal ortholog sequences. These assays are enabling more informative and efficient preclinical gene therapy development and contributing critical data in IND-enabling studies

Learning Objectives:
  • Simultaneous AAV vector biodistribution and transgene mRNA expression analysis 
  • Quantifiable, cell-specific measurement of AAV and transgene in multiplexed assays with cell markers 
  • Rapid, visual germline transmission safety assessment in AAV assays on intact gonadal tissues
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