Articles by Natalie Tully in JoVE
Streamlined Single Cell TCR Isolation and Generation of Retroviral Vectors for In Vitro and In Vivo Expression of Human TCRs Maran L. Sprouse1, Gabriele Blahnik2, Thomas Lee1, Natalie Tully1, Pinaki Benarjee3, Eddie A. James2, Maria J. Redondo4, Matthew L. Bettini1, Maria Bettini1 1 The current protocol combines single cell paired human TCR alpha and beta chain sequencing with streamlined generation of retroviral vectors compatible with in vitro and in vivo TCR expression.
Other articles by Natalie Tully on PubMed
Rapid Identification and Expression of Human TCRs in Retrogenic Mice Journal of Immunological Methods. Dec, 2016 | Pubmed ID: 27589924 Single-cell paired TCR identification is a powerful tool, but has been limited in its previous incompatibility with further functional analysis. The current protocol describes a method to clone and functionally evaluate in vivo TCRs derived from single antigen-responsive human T cells and monoclonal T cell lines. We have improved upon current PCR-based TCR sequencing protocols by developing primers that allow amplification of human TCRα and TCRβ variable regions, while incorporating specific restriction cut sites for direct subcloning into the template retroviral vector. This streamlined approach for generating human:mouse chimeric TCR vectors allows for rapid TCR expression in humanized-retrogenic (hu-Rg) mice through retroviral mediated stem cell gene transfer. Using widely available techniques and equipment, this method is easily adaptable by most laboratories. This is the first TCR identification protocol that is efficiently combined with subsequent in vivo TCR expression.