title: Establishment and validation of analytical reference panels for the standardization of quantitative BCR-ABL1 measurements on the international scale.
abstract: BACKGROUND
  Current guidelines for managing Philadelphia-positive chronic myeloid leukemia include monitoring the expression of the BCR-ABL1 (breakpoint cluster region/c-abl oncogene 1, non-receptor tyrosine kinase) fusion gene by quantitative reverse-transcription PCR (RT-qPCR). Our goal was to establish and validate reference panels to mitigate the interlaboratory imprecision of quantitative BCR-ABL1 measurements and to facilitate global standardization on the international scale (IS).
  
  
  METHODS
  Four-level secondary reference panels were manufactured under controlled and validated processes with synthetic Armored RNA Quant molecules (Asuragen) calibrated to reference standards from the WHO and the NIST. Performance was evaluated in IS reference laboratories and with non-IS-standardized RT-qPCR methods.
  
  
  RESULTS
  For most methods, percent ratios for BCR-ABL1 e13a2 and e14a2 relative to ABL1 or BCR were robust at 4 different levels and linear over 3 logarithms, from 10% to 0.01% on the IS. The intraassay and interassay imprecision was <2-fold overall. Performance was stable across 3 consecutive lots, in multiple laboratories, and over a period of 18 months to date. International field trials demonstrated the commutability of the reagents and their accurate alignment to the IS within the intra- and interlaboratory imprecision of IS-standardized methods.
  
  
  CONCLUSIONS
  The synthetic calibrator panels are robust, reproducibly manufactured, analytically calibrated to the WHO primary standards, and compatible with most BCR-ABL1 RT-qPCR assay designs. The broad availability of secondary reference reagents will further facilitate interlaboratory comparative studies and independent quality assessment programs, which are of paramount importance for worldwide standardization of BCR-ABL1 monitoring results and the optimization of current and new therapeutic approaches for chronic myeloid leukemia.
authors.affiliation: National Genetics Reference Laboratory Wessex, Salisbury District Hospital, Salisbury, UK.
authors.forename: Helen E
authors.initials: HE
authors.lastname: White
authors.forename: John
authors.initials: J
authors.lastname: Hedges
authors.forename: Israel
authors.initials: I
authors.lastname: Bendit
authors.forename: Susan
authors.initials: S
authors.lastname: Branford
authors.forename: Dolors
authors.initials: D
authors.lastname: Colomer
authors.forename: Andreas
authors.initials: A
authors.lastname: Hochhaus
authors.forename: Timothy
authors.initials: T
authors.lastname: Hughes
authors.forename: Suzanne
authors.initials: S
authors.lastname: Kamel-Reid
authors.forename: Dong-Wook
authors.initials: DW
authors.lastname: Kim
authors.forename: Vijay
authors.initials: V
authors.lastname: Modur
authors.forename: Martin C
authors.initials: MC
authors.lastname: Müller
authors.forename: Katia B
authors.initials: KB
authors.lastname: Pagnano
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Pane
authors.forename: Jerry
authors.initials: J
authors.lastname: Radich
authors.forename: Nicholas C P
authors.initials: NC
authors.lastname: Cross
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Labourier
mesh_terms: D016313:Genes, abl; D005820:Genetic Testing / Q000379:methods*; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000175:diagnosis / Q000235:genetics* / Q000503:physiopathology; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000592:standards*; D012720:Severity of Illness Index