Solid-phase synthesis of a biotin derivative and its application to the development of anti-biotin antibodies.
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 N(epsilon)-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.
Papasarantos
Ilias
I
Immunopeptide Chemistry Lab, Institute of Radioisotopes & Radiodiagnostic Products, National Centre for Scientific Research Demokritos, Patriarchou Grigoriou & Neapoleos, Aghia Paraskevi Attikis, Athens 153 10, Greece.
Klimentzou
Persefoni
P
Koutrafouri
Vassiliki
V
Anagnostouli
Maria
M
Zikos
Christos
C
Paravatou-Petsotas
Maria
M
Livaniou
Evangelia
E
D000818:Animals; D000906:Antibodies / Q000032:analysis / Q000276:immunology*; D001710:Biotin / Q000031:analogs & derivatives* / Q000138:chemical synthesis* / Q000737:chemistry / Q000276:immunology*; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D006367:HeLa Cells; D006801:Humans; D007120:Immunochemistry; D008239:Lysine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000276:immunology; D019809:Streptavidin / Q000032:analysis
D000906:Antibodies; D001710:Biotin; D019809:Streptavidin; D008239:Lysine