title: Influence of storage conditions and extraction methods on the quantity and quality of circulating cell-free DNA (ccfDNA): the SPIDIA-DNAplas External Quality Assessment experience.
abstract: BACKGROUND
  Circulating cell-free DNA (ccfDNA) has been confirmed as a useful biomarker in cancer and pre-natal clinical practice. One of the main critical points in using ccfDNA is a lack of standardisation for sample processing methods, storage conditions, procedures for extraction, and quantification that can affect ccfDNA quality and quantity. We report the results obtained from the SPIDIA-DNAplas, one of the EU SPIDIA (Standardisation and improvement of generic pre-analytical tools and procedures for in vitro diagnostics) subprojects based on the implementation of an External Quality Assessment scheme for the evaluation of the influence of the pre-analytical phase on ccfDNA. This is the first reported quality control scheme targeting ccfDNA for pre-analytical phase studies.
  
  
  METHODS
  Fifty-six laboratories throughout Europe were recruited. The participating laboratories received the same plasma sample and extracted ccfDNA by using their own procedures, at defined plasma storage conditions, and sent the isolated ccfDNA to the SPIDIA facility for analyses. Laboratory performance was evaluated by using specific quality parameters such as ccfDNA integrity (by multiplex PCR) and yield (by qPCR).
  
  
  RESULTS
  The analysis of the ccfDNA extracted by the laboratories showed that most of them (53 of 56) were able to recover ccfDNA but only 12.5% recovered non-fragmented ccfDNA. Extraction methods specifically designed for ccfDNA preserved the integrity profile.
  
  
  CONCLUSIONS
  The evidence-based results of the SPIDIA-DNAplas EQA have been proposed as a basis for the development of a Technical Specification by the European Committee for standardisation (CEN).
authors.forename: Francesca
authors.initials: F
authors.lastname: Malentacchi
authors.forename: Sara
authors.initials: S
authors.lastname: Pizzamiglio
authors.forename: Paolo
authors.initials: P
authors.lastname: Verderio
authors.forename: Mario
authors.initials: M
authors.lastname: Pazzagli
authors.forename: Claudio
authors.initials: C
authors.lastname: Orlando
authors.forename: Chiara Maura
authors.initials: CM
authors.lastname: Ciniselli
authors.forename: Kalle
authors.initials: K
authors.lastname: Günther
authors.forename: Stefania
authors.initials: S
authors.lastname: Gelmini
mesh_terms: D001793:Blood Preservation / Q000379:methods* / Q000592:standards*; D001800:Blood Specimen Collection / Q000379:methods* / Q000592:standards*; D004247:DNA / Q000097:blood* / Q000302:isolation & purification* / Q000592:standards; D006801:Humans; D011786:Quality Control
chemical_list: D004247:DNA