title: Species identification in cell culture: a two-pronged molecular approach.
abstract: Species identification of cell lines and detection of cross-contamination are crucial for scientific research accuracy and reproducibility. Whereas short tandem repeat profiling offers a solution for a limited number of species, primarily human and mouse, the standard method for species identification of cell lines is enzyme polymorphism. Isoezymology, however, has its own drawbacks; it is cumbersome and the data interpretation is often difficult. Furthermore, the detection sensitivity for cross-contamination is low; it requires large amounts of the contaminant present and cross-contamination within closely related species may go undetected. In this paper, we describe a two-pronged molecular approach that addresses these issues by targeting the mitochondrial genome. First, we developed a multiplex PCR-based assay to rapidly identify the most common cell culture species and quickly detect cross-contaminations among these species. Second, for speciation and identification of a wider variety of cell lines, we amplified and sequenced a 648-bp region, often described as the "barcode region" by using a universal primer mix targeted at conserved sequences of the cytochrome C oxidase I gene (COI). This method was challenged with a panel of 67 cell lines from 45 diverse species. Implementation of these assays will accurately determine the species of cell lines and will reduce the problems of misidentification and cross-contamination that plague research efforts.
authors.affiliation: ATCC (American Type Culture Collection), 10801 University Boulevard, Manassas, VA 20110, USA.
authors.forename: Jason K
authors.initials: JK
authors.lastname: Cooper
authors.forename: Greg
authors.initials: G
authors.lastname: Sykes
authors.forename: Steve
authors.initials: S
authors.lastname: King
authors.forename: Karin
authors.initials: K
authors.lastname: Cottrill
authors.forename: Natalia V
authors.initials: NV
authors.lastname: Ivanova
authors.forename: Robert
authors.initials: R
authors.lastname: Hanner
authors.forename: Pranvera
authors.initials: P
authors.lastname: Ikonomi
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D003576:Electron Transport Complex IV / Q000378:metabolism; D006801:Humans; D020014:K562 Cells; D010802:Phylogeny; D021122:Protein Subunits / Q000378:metabolism; D013045:Species Specificity
chemical_list: D021122:Protein Subunits; D003576:Electron Transport Complex IV