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A method for the quantification of intracellular zidovudine nucleotides


Kuster, H; Vogt, M; Joos, B; Nadai, V; Luthy, R (1991). A method for the quantification of intracellular zidovudine nucleotides. Journal of Infectious Diseases, 164(4):773-776.

Abstract

An assay to quantify the phosphorylation products of zidovudine (AZT) in peripheral blood mononuclear cells (PBMC) was developed. Extracts of PBMC were separated by high-performance liquid chromatography. Eluted AZT mono- (MP), di- (DP), and triphosphate (TP) were collected in separate portions. Treatment with alkaline phosphatase yielded equimolar amounts ofAZT, which after solid-phase enrichment were assayed by radioimmunoassay. Detection limit was 0.1 pmol/106 PBMC for each nucleotide. Recoveries of 102%-118% were observed. AZT nucleotides were measured in samples from three patients receiving 250 mg ofAZT every 12 h. Intracellular concentrations of AZT-MP after 1-2 h ranged from 0.9 to 1.4 pmol/106 PBMC and then declined to 0.3-1.1 pmol/106 PBMC after 4 h. AZT-DP and AZT-TP reached concentrations of 0.3-0.5 pmol/106 PBMC after 1-2 h and could not be detected after 4 h in any of the three patients. Duplicate determinations deviated by <20%

Abstract

An assay to quantify the phosphorylation products of zidovudine (AZT) in peripheral blood mononuclear cells (PBMC) was developed. Extracts of PBMC were separated by high-performance liquid chromatography. Eluted AZT mono- (MP), di- (DP), and triphosphate (TP) were collected in separate portions. Treatment with alkaline phosphatase yielded equimolar amounts ofAZT, which after solid-phase enrichment were assayed by radioimmunoassay. Detection limit was 0.1 pmol/106 PBMC for each nucleotide. Recoveries of 102%-118% were observed. AZT nucleotides were measured in samples from three patients receiving 250 mg ofAZT every 12 h. Intracellular concentrations of AZT-MP after 1-2 h ranged from 0.9 to 1.4 pmol/106 PBMC and then declined to 0.3-1.1 pmol/106 PBMC after 4 h. AZT-DP and AZT-TP reached concentrations of 0.3-0.5 pmol/106 PBMC after 1-2 h and could not be detected after 4 h in any of the three patients. Duplicate determinations deviated by <20%

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:National licences > 142-005
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Health Sciences > Immunology and Allergy
Health Sciences > Infectious Diseases
Language:English
Date:1 October 1991
Deposited On:09 Oct 2018 13:59
Last Modified:15 Apr 2021 14:47
Publisher:Oxford University Press
ISSN:0022-1899
OA Status:Green
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1093/infdis/164.4.773
PubMed ID:1894938

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