title: Detection of cell-free RNA in children with neuroblastoma and comparison with that of whole blood cell RNA.
abstract: BACKGROUND
  Since there is no validated assay to monitor disease in children with neuroblastoma (NB), we tested whether NB specific cell-free RNA could be detected in their plasma samples. Moreover, with the aim of reducing patients' discomfort, we compared this assay to a recently standardized procedure that uses a larger amount of whole blood.
  
  
  METHODS
  Using conditions that excluded RNA recovery from contaminating tumor cells, the total amount of cell-free RNA present in healthy children and patients with NB was quantified. Expression of tyrosine hydroxylase (TH) was assayed by quantitative RT-PCR.
  
  
  RESULTS
  In patients with NB the amount of cell-free RNA was higher than in healthy children. However, it was less and more degraded than in healthy adults. The median amount of cell-free RNA that was reverse transcribed, measured through the use of standard curves for reference genes, was 0.03 (range 0-30) pg of input RNA, that is, always less than 1/10,000 of that reverse transcribed from total RNA extracted from whole cells. Despite the presence of disease and the positive results obtained with RNA extracted from peripheral blood cells, few cell-free RNA samples tested positive by the TH assay. Similar results were obtained also with TH primers specifically designed to amplify 50 bp RNA fragments.
  
  
  CONCLUSIONS
  These findings suggest that for monitoring disease status detection of cell-free tumor-specific RNAs in patients with NB is not a reliable alternative to whole cell RNA.
authors.affiliation: Laboratory of Oncology, Gaslini Institute, Genoa, Italy. mariavaleriacorrias@ospedale-gaslini.ge.it
authors.forename: Maria Valeria
authors.initials: MV
authors.lastname: Corrias
authors.forename: Angela
authors.initials: A
authors.lastname: Pistorio
authors.forename: Giuliana
authors.initials: G
authors.lastname: Cangemi
authors.forename: Gino
authors.initials: G
authors.lastname: Tripodi
authors.forename: Barbara
authors.initials: B
authors.lastname: Carlini
authors.forename: Paola
authors.initials: P
authors.lastname: Scaruffi
authors.forename: Paolo
authors.initials: P
authors.lastname: Fardin
authors.forename: Alberto
authors.initials: A
authors.lastname: Garaventa
authors.forename: Vito
authors.initials: V
authors.lastname: Pistoia
authors.forename: Riccardo
authors.initials: R
authors.lastname: Haupt
mesh_terms: D000293:Adolescent; D000328:Adult; D014408:Biomarkers, Tumor / Q000097:blood* / Q000235:genetics; D001773:Blood Cells / Q000737:chemistry; D001932:Brain Neoplasms / Q000175:diagnosis*; D002648:Child; D002675:Child, Preschool; D019075:Electrophoresis, Capillary; D005260:Female; D006403:Hematologic Tests / Q000379:methods*; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008297:Male; D009447:Neuroblastoma / Q000175:diagnosis*; D012313:RNA / Q000032:analysis / Q000097:blood*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D014408:Biomarkers, Tumor; D012313:RNA