Use of l-Lysine Fluorescence Derivatives as Tracers To Enhance the Performance of Polarization Fluoroimmunoassays. A Study Using Two Herbicides as Model Antigens

Fluorescence polarization immunoassay (FPIA) is a convenient homogeneous assay, the use of which is restricted in environmental analysis by low sensitivity and matrix effects. We selected the herbicides 2,4D and 2,4,5T to synthesize new l-lysine-based fluorescent tracers using solid-phase chemistry....

Full description

Saved in:
Bibliographic Details
Published in:Analytical chemistry (Washington) 2002-06, Vol.74 (11), p.2513-2521
Main Authors: Hatzidakis, George I, Tsatsakis, Aristidis M, Krambovitis, Elias K, Spyros, Apostolos, Eremin, Sergei A
Format: Article
Language:English
Subjects:
2,4,5-Trichlorophenoxyacetic Acid - analysis
2,4-Dichlorophenoxyacetic Acid - analysis
Animals
Antigens - analysis
Chemistry
Fluorescence
Fluorescence Polarization Immunoassay - methods
Fluorescent Dyes - chemistry
Herbicides
Herbicides - analysis
Indicators and Reagents
Lysine - chemistry
Rabbits
Serum Albumin, Bovine
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Fluorescence polarization immunoassay (FPIA) is a convenient homogeneous assay, the use of which is restricted in environmental analysis by low sensitivity and matrix effects. We selected the herbicides 2,4D and 2,4,5T to synthesize new l-lysine-based fluorescent tracers using solid-phase chemistry. In addition, three different immunogens of 2,4,5T were prepared for immunization and antibody production. The new tracers and antibodies were adapted to FPIA. Tracers with the hapten attached to the α-aminogroup of l-lysine and fluorescein to the e-amino group exhibited at least a 5-fold increased sensitivity when compared to the previously reported ethylenediamine-based tracer (2,4D-EDA-F). The isomeric structure (hapten attached to the e-amino and fluorescein to the a-amino group) appeared 7.6 times less sensitive, and all other alternative structures exhibited even lower sensitivities. This observation was confirmed against the monoclonal anti-2,4D antibody E2/G2 and polyclonal anti-2,4,5T antibodies. The affinity constant of 2,4D-EDA-F with E2/G2 was 8.1 times higher when compared with the new tracer, suggesting the more specific nature of the l-lysine-based tracer, the use of which leads to a more sensitive assay. This type of tracer could improve performance and lower substantially the detection limits of FPIAs.
ISSN:0003-2700
1520-6882
DOI:10.1021/ac011051x