title: Comparison of lentiviral vector titration methods.
abstract: BACKGROUND
  Lentiviral vectors are efficient vehicles for stable gene transfer in dividing and non-dividing cells. Several improvements in vector design to increase biosafety and transgene expression, have led to the approval of these vectors for use in clinical studies. Methods are required to analyze the quality of lentiviral vector production, the efficiency of gene transfer and the extent of therapeutic gene expression.
  
  
  RESULTS
  We compared lentiviral vector titration methods that measure pg p24/ml, RNA equivalents/ml, transducing units (TU/ml) or mRNA equivalents. The amount of genomic RNA in vector particles proves to be reliable to assess the production quality of vectors encoding non-fluorescent proteins. However, the RNA and p24 titers of concentrated vectors are rather poor in predicting transduction efficiency, due to the high variability of vector production based on transient transfection. Moreover, we demonstrate that transgenic mRNA levels correlate well with TU and can be used for functional titration of non-fluorescent transgenes.
  
  
  CONCLUSIONS
  The different titration methods have specific advantages and disadvantages. Depending on the experimental set-up one titration method should be preferred over the others.
authors.affiliation: Laboratory for Molecular Virology and Gene Therapy, K.U.Leuven and IRC KULAK, Flanders, Belgium. martine.geraerts@med.kuleuven.be
authors.forename: Martine
authors.initials: M
authors.lastname: Geraerts
authors.forename: Sofie
authors.initials: S
authors.lastname: Willems
authors.forename: Veerle
authors.initials: V
authors.lastname: Baekelandt
authors.forename: Zeger
authors.initials: Z
authors.lastname: Debyser
authors.forename: Rik
authors.initials: R
authors.lastname: Gijsbers
mesh_terms: D002874:Chromosome Mapping / Q000379:methods; D004279:DNA, Viral / Q000032:analysis*; D005822:Genetic Vectors / Q000235:genetics*; D016679:Genome, Viral / Q000235:genetics; D016086:Lentivirus / Q000235:genetics* / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000032:analysis*; D014162:Transfection / Q000379:methods*
chemical_list: D004279:DNA, Viral; D012367:RNA, Viral