title: [A method to detect gene transfection efficacy by flow cytometry].
abstract: OBJECTIVE
  Analysis of gene transfer and expression is conventionally inferred from the percentage of positive cells expressing reporter gene in total cells, referred as transfection rate, by investigators counting under a microscope or fluoroscope, which was called as manual counting. But in many cases, it is not accurate and easily influenced by the subjectivity of observer. This study was designed to seek a convenient method to assess objectively and accurately the efficacy of gene transfer and expression.
  
  
  METHODS
  Hepatocellular carcinoma(HCC) HepG2 cells were infected with a recombinant adenovirus expressing green fluorescent protein(AdCMV/GFP) at a series of multiplicities of infection(MOIs). 24 h later, the transfection rates were assessed by manual counting under fluorescent microscope. Meanwhile, besides transfection rates, fluorescent indices(FIs) which indicated the efficiency of gene transfer and expression were analyzed by flow cytometry (FCM). Transfection efficiencies of AdCMV/GFP to HCC Hep3B, Bel7402, SMMC7721 cells and nasopharyngeal carcinoma CNE-2 cells were also tested by FCM.
  
  
  RESULTS
  Although transfection rates by FCM were slightly higher than that by manual counting, both were logarithmic correlative with vector doses. The stirring was that FIs by FCM showed compellent linear correlation with vector doses (r = 0.9984, P < 0.001). The efficiency of gene transfer in other cells by FCM were similar to that in HepG2.
  
  
  CONCLUSIONS
  The efficiency of gene transfer and expression in mammalian cells can be easily analyzed by flow cytometry, which is more sensitive, objective, and accurate than manual counting, especially in assessing the efficiency of multiple gene transfer (multi-copies per cell) and expression.
authors.affiliation: Biotherapy Center, Cancer Center, Sun Yat-sen University, Guangzhou 510060, P. R. China.
authors.forename: Ran-yi
authors.initials: RY
authors.lastname: Liu
authors.forename: Hui-ling
authors.initials: HL
authors.lastname: Luo
authors.forename: Hai-lin
authors.initials: HL
authors.lastname: Feng
authors.forename: Ji-lin
authors.initials: JL
authors.lastname: Peng
authors.forename: Ti-yu
authors.initials: TY
authors.lastname: Cai
authors.forename: Wen-lin
authors.initials: WL
authors.lastname: Huang
authors.forename: Yi-xin
authors.initials: YX
authors.lastname: Zeng
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D005434:Flow Cytometry / Q000379:methods*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured