title: Multicenter evaluation of a novel nanoparticle immunoassay for 5-fluorouracil on the Olympus AU400 analyzer.
abstract: BACKGROUND
  5-Fluorouracil (5-FU) is the most widely used chemotherapy drug, primarily against gastrointestinal, head and neck, and breast cancers. 5-FU has large pharmacokinetic variability resulting in unexpected toxicity or ineffective treatment. Therapeutic drug management of 5-FU minimizes toxicity and improves outcome. A nanoparticle-based immunoassay was developed to provide oncologists with a rapid, cost-effective tool for determining 5-FU plasma concentrations.
  
  
  METHODS
  Monoclonal antibodies, bound to nanoparticles, were used to develop an immunoassay for the Olympus AU400. Assay precision, linearity, calibration stability, and limit of detection were run at multiple centers; interference, cross-reactivity, lower limit of quantitation and recovery at 1 center. Clinical samples collected from 4 cancer centers were analyzed for 5-FU concentrations by liquid chromatography-tandem mass spectrometry and compared with the immunoassay results.
  
  
  RESULTS
  With calibrators from 0 to 1800 ng/mL 5-FU and autodilution, concentrations up to 9000 ng/mL could be determined. Time to first result was 10 minutes, and 400 samples per hour could be quantitated from a standard curve stored for >30 days. Imprecision across all laboratories was <5%, and the assay was linear upon dilution over the entire range. Cross-reactivities for dihydro-5-FU, uracil, capecitabine, and tegafur were <1%, 9.9%, 0.05%, and 0.23%, respectively. The limit of detection was 52 ng/mL with a lower limit of quantitation of 86 ng/mL. Assay results of clinical samples (93-1774 ng/mL) correlated with liquid chromatography-tandem mass spectrometry results: (R = 0.9860, slope 1.035, intercept 10.87 ng/mL).
  
  
  CONCLUSIONS
  This novel immunoassay is suitable for quantitating 5-FU plasma concentrations with advantages of speed, small sample size, minimal sample pretreatment, and application on automated instrumentation. These advantages enable efficient therapeutic drug management of 5-FU in clinical practice.
authors.affiliation: University of Pittsburgh Cancer Institute, Pittsburgh, Pennsylvania, USA.
authors.forename: Jan H
authors.initials: JH
authors.lastname: Beumer
authors.forename: M
authors.initials: M
authors.lastname: Boisdron-Celle
authors.forename: William
authors.initials: W
authors.lastname: Clarke
authors.forename: Jodi B
authors.initials: JB
authors.lastname: Courtney
authors.forename: Merrill J
authors.initials: MJ
authors.lastname: Egorin
authors.forename: Erick
authors.initials: E
authors.lastname: Gamelin
authors.forename: Rebecca L
authors.initials: RL
authors.lastname: Harney
authors.forename: Catherine
authors.initials: C
authors.lastname: Hammett-Stabler
authors.forename: Sandy
authors.initials: S
authors.lastname: Lepp
authors.forename: Yunying
authors.initials: Y
authors.lastname: Li
authors.forename: Gregory D
authors.initials: GD
authors.lastname: Lundell
authors.forename: Gwen
authors.initials: G
authors.lastname: McMillin
authors.forename: Gerard
authors.initials: G
authors.lastname: Milano
authors.forename: Salvatore J
authors.initials: SJ
authors.lastname: Salamone
mesh_terms: D000911:Antibodies, Monoclonal; D002138:Calibration; D016903:Drug Monitoring / Q000379:methods; D005472:Fluorouracil / Q000097:blood*; D006801:Humans; D007118:Immunoassay / Q000191:economics / Q000295:instrumentation / Q000379:methods*; D057230:Limit of Detection; D053758:Nanoparticles; D009391:Nephelometry and Turbidimetry; D015203:Reproducibility of Results; D013997:Time Factors
chemical_list: D000911:Antibodies, Monoclonal; D005472:Fluorouracil