title: Quantum dot labeling and tracking of cultured limbal epithelial cell transplants in vitro.
abstract: OBJECTIVE
  Cultured human limbal epithelial cells (HLECs) have shown promise in the treatment of limbal stem cell deficiency but little is known about their survival, behavior, and long-term fate after transplantation. The aim of this research was to evaluate, in vitro, quantum dot (Qdot) technology as a tool for tracking transplanted HLECs.
  
  
  METHODS
  In vitro cultured HLECs were labeled with Qdot nanocrystals. Toxicity was assessed using live-dead assays. The effect on HLEC function was assessed using colony-forming efficiency assays and expression of CK3, P63alpha, and ABCG2. Sheets of cultured HLECs labeled with Qdot nanocrystals were transplanted onto decellularized human corneoscleral rims in an organ culture model and observed to investigate the behavior of transplanted cells.
  
  
  RESULTS
  Quantum dot labeling had no detrimental effect on HLEC viability or function in vitro. Proliferation resulted in a gradual reduction in Qdot signal but sufficient signal was present to allow tracking of cells through multiple generations. Cells labeled with Qdots could be reliably detected and observed using confocal microscopy for at least 2 weeks after transplantation in our organ culture model. In addition, it was possible to label and observe epithelial cells in intact human corneas by using the Rostock corneal module adapted for use with the Heidelberg HRA.
  
  
  CONCLUSIONS
  This work demonstrates that Qdots combined with existing clinical equipment could be used to track HLEC for up to 2 weeks after transplantation; however, our model does not permit the assessment of cell labeling beyond 2 weeks. Further characterization in in vivo models are required.
authors.affiliation: Department of Ocular Biology and Therapeutics, UCL Institute of Ophthalmology, London, United Kingdom 2National Institute for Health Research Biomedical Research Centre, Moorfields Eye Hospital and UCL Institute of Ophthalmology, London, United Kingdom.
authors.forename: Nuria
authors.initials: N
authors.lastname: Genicio
authors.affiliation: Comprehensive Cancer Imaging Centre, Imperial College, South Kensington, London, London, United Kingdom 4Department of Chemistry, Imperial College London, South Kensington, London, United Kingdom.
authors.forename: Juan
authors.initials: J
authors.lastname: Gallo Paramo
authors.affiliation: Department of Ocular Biology and Therapeutics, UCL Institute of Ophthalmology, London, United Kingdom 2National Institute for Health Research Biomedical Research Centre, Moorfields Eye Hospital and UCL Institute of Ophthalmology, London, United Kingdom 5C.
authors.forename: Alex J
authors.initials: AJ
authors.lastname: Shortt
mesh_terms: D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D017690:Cell Transplantation / Q000379:methods*; D002478:Cells, Cultured; D019573:Epithelium, Corneal / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D066298:In Vitro Techniques; D053558:Keratin-3 / Q000378:metabolism; D016850:Limbus Corneae / Q000166:cytology*; D008854:Microscopy, Electron; D009363:Neoplasm Proteins / Q000378:metabolism; D045663:Quantum Dots / Q000009:adverse effects / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000378:metabolism
chemical_list: C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; C479360:KRT3 protein, human; D053558:Keratin-3; D009363:Neoplasm Proteins; C115051:TP63 protein, human; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins