Development of an offline noncompetitive flow immunoassay for the determination of interleukin-8 in cell samples

Forskningsoutput: TidskriftsbidragArtikel i vetenskaplig tidskrift

Abstract

A noncompetitive flow immunoassay system (FIA) for the analysis of interleukin-8 (IL-8) in cell samples was developed. Affinity interaction assays based on offline incubation of excess labeled antibodies and antigen (IL-8) were carried out. The residual unbound labeled antibody was trapped in an immunoaffinity column with immobilized IL-8 while the immunocomplex, labeled antibody/IL-8, was detected by a fluorescence detector. Two fluorophores, FLUOS and Cy5.5, were conjugated with IL-8 antibody. Optimization and comparison between the two fluorescent labeled antibodies were performed with regard to pH, antibody concentration, flow rate, injection volume, and association time. Additionally, a horseradish peroxidase enzyme label was used for the conjugation to the anti-IL-8. The enzyme substrate reaction was optimized with respect to temperature and length of the substrate reaction coil. The detection limits were found to be 200 amol using the FLUOS-labeled anti-IL-8 and 1 fmol using the Cy5.5 fluorescence label. The developed FIA technique was applied for the analysis of IL-8 in cell samples. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used to identify IL-8 in the cell samples.

Detaljer

Författare
Externa organisationer
  • AstraZeneca
  • Pharmacia & Upjohn
Forskningsområden

Ämnesklassifikation (UKÄ) – OBLIGATORISK

  • Annan medicinsk grundvetenskap

Nyckelord

Originalspråkengelska
Sidor (från-till)46-54
Antal sidor9
TidskriftAnalytical Biochemistry
Volym279
Utgåva nummer1
StatusPublished - 2000 mar 1
PublikationskategoriForskning
Peer review utfördJa
Externt publiceradJa