title: Cytokine marker measurement in human neuroblastoma cells with supersensitive and multiplex assay: MUSTag technology.
abstract: BACKGROUND
  Recently, various sets of protein -biomarkers have been discovered in important diseases such as cancers, brain stroke and heart attack. However, clinical validation is difficult and time-consuming by individual assays or because of very low concentrations at early stages of the diseases. We have developed assay technology through an innovative modification of the immuno-PCR method for the super-sensitive and multiplex detection of target biomarkers.
  
  
  METHODS
  In the assay technology, each different oligo-tag simultaneously detects multiplex protein targets with extremely high-level sensitivity in a dose-dependent manner by qRT-PCR (maximum: three plexes). In this study, we measured specific secreted protein concentrations in the culture supernatant of a 24-h culture of transfected SH-SY5Y cells with MUSTag.
  
  
  RESULTS
  There was a significant increase in the protein level of tumor necrosis factor (TNF)-α measured with extremely high-level sensitivity (≥10 pg/mL). Compared with negative controls, the levels of TNF-α increased from 16.9 to 28.1 pg/mL (p = 0.011).
  
  
  CONCLUSIONS
  We suggest that our assay technology might be of clinical value in treating patients with cancer, cerebral ischemia, or patients who need a prompt and predictive diagnosis for adequate treatment.
authors.affiliation: Department of Anesthesiology, Tokyo Medical University, Tokyo, Japan. miyamiya717@hotmail.co.jp
authors.forename: Ryoichi
authors.initials: R
authors.lastname: Miyashita
authors.forename: Li
authors.initials: L
authors.lastname: Chen
authors.forename: Yoshihito
authors.initials: Y
authors.lastname: Morizane
authors.forename: Yuji
authors.initials: Y
authors.lastname: Takeshita
authors.forename: Hidekimi
authors.initials: H
authors.lastname: Fukui
authors.forename: Taku
authors.initials: T
authors.lastname: Ishizaki
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Hatakeyama
authors.forename: Kiyoshige
authors.initials: K
authors.lastname: Ohseto
authors.forename: Futoshi
authors.initials: F
authors.lastname: Shibasaki
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Uchino
mesh_terms: D042683:Angiopoietin-1 / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D039621:Eukaryotic Initiation Factor-3 / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D009447:Neuroblastoma; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis*
chemical_list: D042683:Angiopoietin-1; D039621:Eukaryotic Initiation Factor-3; D034741:RNA, Small Interfering; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha; D049452:Green Fluorescent Proteins