title: Quality assessment of human mitochondrial DNA quantification: MITONAUTS, an international multicentre survey.
abstract: Mitochondrial DNA quantification by qPCR is used in the context of many diseases and toxicity studies but comparison of results between laboratories is challenging. Through two multigroup distributions of DNA samples from human cell lines, the MITONAUTS group anonymously compared mtDNA/nDNA quantification across nine laboratories involved in HIV research worldwide. Eight of the nine sites showed significant correlation between them (mean raw data R(2)=0.664; log(10)-transformed data R(2)=0.844). Although mtDNA/nDNA values were well correlated between sites, the inter-site variability on the absolute measurements remained high with a mean (range) coefficient of variation of 71 (37-212) %. Some variability appeared cell line-specific, probably due to chromosomal alterations or pseudogenes affecting the quantification of certain genes, while within cell line variability was likely due to differences in calibration of the standard curves. The use of two mtDNA and two single copy nDNA genes with highly specific primers to quantify each genome would help address copy number variants. Our results indicate that sample shipment must be done frozen and that absolute mtDNA/nDNA ratio values cannot readily be compared between laboratories, especially if assessing cultured cell mtDNA content. However, within laboratory and relative mtDNA/nDNA comparisons between laboratories should be reliable.
authors.affiliation: Department of Pathology & Laboratory Medicine, University of British Columbia, Vancouver, Canada. helene.cote@ubc.ca
authors.forename: Hélène C F
authors.initials: HC
authors.lastname: Côté
authors.forename: Mariana
authors.initials: M
authors.lastname: Gerschenson
authors.forename: Ulrich A
authors.initials: UA
authors.lastname: Walker
authors.forename: Oscar
authors.initials: O
authors.lastname: Miro
authors.forename: Gloria
authors.initials: G
authors.lastname: Garrabou
authors.forename: Emma
authors.initials: E
authors.lastname: Hammond
authors.forename: Joan
authors.initials: J
authors.lastname: Villarroya
authors.forename: Marta
authors.initials: M
authors.lastname: Giralt
authors.forename: Francesc
authors.initials: F
authors.lastname: Villarroya
authors.forename: Paola
authors.initials: P
authors.lastname: Cinque
authors.forename: Elena
authors.initials: E
authors.lastname: Garcia-Arumi
authors.forename: Antonio L
authors.initials: AL
authors.lastname: Andreu
authors.forename: Marcello
authors.initials: M
authors.lastname: Pinti
authors.forename: Andrea
authors.initials: A
authors.lastname: Cossarizza
mesh_terms: D002460:Cell Line; D004272:DNA, Mitochondrial / Q000032:analysis* / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D057089:Pathology, Molecular / Q000379:methods* / Q000592:standards*; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards*; D015203:Reproducibility of Results; D013048:Specimen Handling / Q000379:methods
chemical_list: D004272:DNA, Mitochondrial