Solid-Phase Synthesis of a Biotin Derivative and its Application to the Development of Anti-Biotin Antibodies |
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Authors: | Ilias Papasarantos Persefoni Klimentzou Vassiliki Koutrafouri Maria Anagnostouli Christos Zikos Maria Paravatou-Petsotas Evangelia Livaniou |
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Affiliation: | 1.Immunopeptide Chemistry Lab, Institute of Radioisotopes & Radiodiagnostic Products,National Centre for Scientific Research “Demokritos”,Athens,Greece;2.Department of Neurology, Medical School,University of Athens, Eginition Hospital,Athens,Greece |
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Abstract: | A biotin derivative, namely biotin–aminocaproic acid–lysine (BAL), was synthesized with solid-phase chemistry, conjugated
to a carrier-protein, and used for rabbit immunization. The aminocaproic acid–lysine “long-arm” was used in order to project
the biotin-hapten above the carrier-protein surface. Lysine was selected due to its Nε-amino group, through which BAL was conjugated to the carrier-protein. BAL was synthesized on a commercially available resin
with the Fmoc-solid-phase strategy; this has simplified the experimental procedure, overcome the need for intermediate purification
steps, and led to a final product of high purity, with high yield. The anti-BAL antibodies recognized free biotin, as shown
with an in-house-developed ELISA, in which biotin conjugated to a synthetic “lysine–dendrimer” was used to coat the ELISA
microwells. In immunocytology and Western-blot experiments, the anti-BAL antibodies led to similar results with those obtained
with streptavidin. Synthetic derivatives of hapten molecules that can be easily prepared with solid-phase chemistry, such
as BAL, may be used for the development of specific antibodies for the corresponding hapten. |
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