Enzyme immunoassay of vitamin d3 metabolites
https://doi.org/10.29235/1561-8331-2020-56-2-197-205
Abstract
We report herein improved version of the synthesis of hapten based on cholecalciferol and its active metabolite 25-hydroxycholecalcalferol. The methodology of obtaining high-molecular immunogenic conjugates of vitamin D3 derivatives with bovine serum albumin and horseradish peroxidase conjugates for direct ELISA was optimized. Immunisation of rabbits was carried out and polyclonal antibodies to 25-hydroxycholecalceferol were obtained and tested in an enzyme-linked immunosorbent assay. To improve the accuracy of the method, the sample preparation procedure was optimized, including the release of vitamin D3 and its active metabolites from complexes with vitamin D-binding protein.
Keywords
25-hydroxycholecalciferol,
vitamin D3,
bioconjugation,
enzyme immunoassay,
vitamin D-binding protein
Supporting agencies: We thank Dr. Cristhop Weise (Freie Universität Berlin) for assistance in obtaining and interpreting the mass-spectra.
About the Authors
A. S. Kruhlik
Institute of Physical Organic Chemistry of the National Academy of Sciences of Belarus
Belarus
Belarus
Aliaksandr S. Kruhlik – Researcher
13, Surganov Str., 220072, Minsk
D. V. Tsaulouski
Institute of Physical Organic Chemistry of the National Academy of Sciences of Belarus
Belarus
Belarus
Dzmitry V. Tsaulouski – Researcher
13, Surganov Str., 220072, Minsk
T. V. Mirantsova
Hemma-test Joint LLC
Belarus
Tatsiana V. Mirantsova – Researcher
Belarus
Tatsiana V. Mirantsova – Researcher
O. L. Sharko
Institute of Physical Organic Chemistry of the National Academy of Sciences of Belarus
Belarus
Belarus
Olga L. Sharko – Ph. D. (Chemistry), Leading Researcher
13, Surganov Str., 220072, Minsk
V. V. Shmanai
Institute of Physical Organic Chemistry of the National Academy of Sciences of Belarus
Belarus
Belarus
Vadim V. Shmanai – Ph. D. (Chemistry), Head of the Laboratory
13, Surganov Str., 220072, Minsk
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