Quantitation of 3'-amino-3'-deoxythymidine (AMT), a toxic catabolite of 3'-azido-3'-deoxythymidine (AZT), by competitive ELISA.
In the present study, a competitive ELISA technique was developed to specifically quantitate 3'-amino-3'-deoxythymidine (AMT), a toxic catabolite of 3'-azido-3'-deoxythymidine (AZT) detected in serum from AZT-treated patients. In order to eliminate cross-reacting AZT, serum sample was extracted with ethylacetate and then AMT was acetylated (Ac-AMT). A 5'-hemisuccinate-AMT-horseradish peroxidase conjugate was used as a tracer in the presence of anti-AMT rabbit antibodies which were raised against a 5' hemisuccinate-AMT-bovine serum albumin immunogen. Bound/free separation was achieved with an anti-rabbit IgG mouse monoclonal antibody insolubilized onto a microtiter plate. The limit of quantification of Ac-AMT was as low as 0.4 ng/ml in serum samples. This ELISA technique was applied for monitoring AMT plasma levels in patients receiving AZT therapy. The intra and inter-individual variations of the AZT/AMT plasma concentration ratios underlined the need for such a specific test in studying the formation of this toxic catabolite.
Ferrua
B
B
Laboratoire de Pharmacologie Expérimentale et Clinique, Faculté de Médecine, Nice, France.
Chakboub
H
H
Roptin
C
C
Garraffo
R
R
Faraj
A
A
Grassi
J
J
Guedj
R
R
Sommadossi
J P
JP
D001667:Binding, Competitive / Q000276:immunology; D015224:Dideoxynucleosides / Q000032:analysis* / Q000276:immunology* / Q000633:toxicity; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D015215:Zidovudine / Q000378:metabolism* / Q000633:toxicity
D015224:Dideoxynucleosides; D015215:Zidovudine; C019252:3'-aminothymidine