Highly sensitive avidin-biotin ELISA for detection of nandrolone and testosterone in dietary supplements
Language English Country Great Britain, England Media print-electronic
Document type Evaluation Study, Journal Article
PubMed
27367148
DOI
10.1002/dta.2005
Knihovny.cz E-resources
- Keywords
- ELISA, anabolics, avidin, biotin, dietary supplements, nandrolone, testosterone,
- MeSH
- Anabolic Agents analysis MeSH
- Avidin chemistry MeSH
- Biotin chemistry MeSH
- Enzyme-Linked Immunosorbent Assay methods MeSH
- Rabbits MeSH
- Limit of Detection MeSH
- Nandrolone analysis MeSH
- Dietary Supplements analysis MeSH
- Testosterone analysis MeSH
- Animals MeSH
- Check Tag
- Rabbits MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
- Names of Substances
- Anabolic Agents MeSH
- Avidin MeSH
- Biotin MeSH
- Nandrolone MeSH
- Testosterone MeSH
Avidin-biotin technology was used for the implementation of an enzyme-linked immunosorbent assay (AB-ELISA) as a sensitive method for the detection of anabolic androgenic steroids (AAS) present in dietary supplements. Using click chemistry, novel haptens (linker-optimized biotinylated nandrolone (NT) and testosterone (T) at positions C-3 and C-17, respectively) were designed and synthesized to be then applied as four different immobilized competitors in a proposed set of four indirect competitive AB-ELISAs. Four rabbit polyclonal antibodies of various specificities were prepared using four different immunogens synthesized from C-3 and C-17 carboxymethyloxime and hemisuccinate derivatives of NT and T, respectively. Assembled AB-ELISAs were characterized to establish method parameters such as a half-maximum inhibition concentration (0.18-12.99 ng/mL), limit of detection (0.004-0.032 ng/mL) and linear working range (the best with 0.02-1.38 ng/mL). The stability of the set simulating storage in different conditions was demonstrated. Cross reactivity (CR) was tested for 59 steroids including both endogenous and synthetic analogues in four assembled AB-systems. The focus was placed on the practical use of the method in detection of various AAS in 49 samples of counterfeit dietary supplements. The concordance between ultra high performance liquid chromatography-mass spectrometry (UHPLC-MS) and the CR corrected data from AB-ELISA indicated the potential of this method even to quantification of T propionate, NT phenyl propionate, and NT decanoate in such a complex matter. Copyright © 2016 John Wiley & Sons, Ltd.
Czech Agriculture and Food Inspection Authority Prague Czech Republic
University of Chemistry and Technology Prague Prague Czech Republic
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