title: Harmonization of BCR-ABL mRNA quantification using a uniform multifunctional control plasmid in 37 international laboratories.
abstract: Individualized PCR strategies hamper comparability of molecular results between different laboratories in several fields of medicine. To harmonize BCR-ABL mRNA quantification an international multicenter trial involving 37 laboratories in 14 countries was initiated using 10 samples, each containing various dilutions (10, 2, 1 and 0.1%) of b3a2 or b2a2 BCR-ABL positive in normal leukocytes and negative controls. A novel control plasmid (pME-2) was designed for external calibration containing BCR-ABL and glucuronidase-beta (GUS) sequences. Median BCR-ABL/ABL ratios were 9.1, 1.8, 0.85 and 0.11% in b3a2 samples and 9.5, 1.6, 0.84 and 0.11% in b2a2 samples. Median BCR-ABL/GUS ratios were 3.4, 0.77, 0.37 and 0.042% in b3a2 samples and 2.8, 0.48, 0.29 and 0.031% in b2a2 samples. The coefficients of variation were 0.62 for ratios BCR-ABL/ABL and 1.03 for ratios BCR-ABL/GUS. Five of 37 evaluable participating laboratories (13%) detected low BCR-ABL copy numbers in negative control samples; one laboratory failed to detect BCR-ABL in a low-level sample. We conclude that the use of a common control plasmid does indeed improve comparability of BCR-ABL mRNA quantification results. However, further standardizing efforts like introducing a calibrator and regular control rounds are needed.
authors.affiliation: III. Medizinische Universitätsklinik, Medizinische Fakultät Mannheim der Universität Heidelberg, Mannheim, Germany. martin.mueller@med3.ma.uni-heidelberg.de
authors.forename: M C
authors.initials: MC
authors.lastname: Müller
authors.forename: P
authors.initials: P
authors.lastname: Erben
authors.forename: G
authors.initials: G
authors.lastname: Saglio
authors.forename: E
authors.initials: E
authors.lastname: Gottardi
authors.forename: C G
authors.initials: CG
authors.lastname: Nyvold
authors.forename: T
authors.initials: T
authors.lastname: Schenk
authors.forename: T
authors.initials: T
authors.lastname: Ernst
authors.forename: S
authors.initials: S
authors.lastname: Lauber
authors.forename: J
authors.initials: J
authors.lastname: Kruth
authors.forename: R
authors.initials: R
authors.lastname: Hehlmann
authors.forename: A
authors.initials: A
authors.lastname: Hochhaus
mesh_terms: D000368:Aged; D005260:Female; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007753:Laboratories; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000097:blood / Q000175:diagnosis / Q000235:genetics*; D008297:Male; D010957:Plasmids; D012333:RNA, Messenger / Q000032:analysis*; D012334:RNA, Neoplasm / Q000235:genetics; D012015:Reference Standards; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000592:standards
chemical_list: D012333:RNA, Messenger; D012334:RNA, Neoplasm; D016044:Fusion Proteins, bcr-abl; D005966:Glucuronidase