title: Efficient and reliable establishment of lymphoblastoid cell lines by Epstein-Barr virus transformation from a limited amount of peripheral blood.
abstract: Lymphoblastoid cell lines (LCLs) transformed by Epstein-Barr virus (EBV) serve as an unlimited resource of human genomic DNA. The protocol that is widely used to establish LCLs involves peripheral blood mononuclear cell isolation by density gradient centrifugation, however, that method requires as much as 5 ml of peripheral blood. In this study, in order to provide a more simple and efficient method for the generation of LCLs, we developed a new protocol using hemolytic reaction to enrich white blood cells for EBV transformation and found that the hemolytic protocol successfully generated LCLs from a small volume (i.e., 0.1 ml) of peripheral blood. To assess the quality of genomic DNA extracted from LCLs established by the hemolytic protocol (LCL-hemolytic), we performed single nucleotide polymorphism (SNP) microarray genotyping using the GeneChip®100 K Array Set (Affymetrix, Inc.). The concordances of the SNP genotyping resulting from genomic DNA from LCL-hemolytic (99.92%) were found to be as good as the technical replicate (99.90%), and Kappa statistics results confirmed the reliability. The findings of this study reveal that the hemolytic protocol is a simple and reliable method for the generation of LCLs, even from a small volume of peripheral blood.
authors.affiliation: Department of Genomic Medical Sciences, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Natsue
authors.initials: N
authors.lastname: Omi
authors.affiliation: Department of Genomic Medical Sciences, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Tokuda
authors.affiliation: Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Ikeda
authors.affiliation: Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Morio
authors.initials: M
authors.lastname: Ueno
authors.affiliation: Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Mori
authors.affiliation: Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Chie
authors.initials: C
authors.lastname: Sotozono
authors.affiliation: Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan.|Department of Frontier Medical Science and Technology for Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kinoshita
authors.affiliation: Department of Genomic Medical Sciences, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Masakazu
authors.initials: M
authors.lastname: Nakano
authors.affiliation: Department of Genomic Medical Sciences, Kyoto Prefectural University of Medicine, Kyoto, Japan.
authors.forename: Kei
authors.initials: K
authors.lastname: Tashiro
mesh_terms: D002461:Cell Line, Transformed; D015894:Genome, Human / Q000235:genetics*; D005838:Genotype; D004854:Herpesvirus 4, Human / Q000502:physiology; D006801:Humans; D007962:Leukocytes / Q000166:cytology / Q000378:metabolism* / Q000821:virology; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000378:metabolism* / Q000821:virology; D008214:Lymphocytes / Q000378:metabolism* / Q000821:virology; D020411:Oligonucleotide Array Sequence Analysis; D020641:Polymorphism, Single Nucleotide; D015203:Reproducibility of Results