PMID: 2120348Oct 4, 1990Paper

A single-step method for the purification of anti-FITC antibodies by use of a coumarin immunosorbent

Journal of Immunological Methods
D Samuel, R A Abuknesha

Abstract

Monoclonal anti-fluorescein isothiocyanate (FITC) antibody cross-reacts with 7-hydroxy coumarin derivatives conjugated to BSA. This property permitted the affinity purification of monoclonal anti-FITC antibodies from ascitic fluid using an-immunosorbent consisting of a 7-hydroxy coumarin derivative linked to Sepharose 4B. Ascitic fluid was applied to the immunosorbent column and, after washing, the bound antibody was eluted under extremely mild conditions using 3 M MgCl2. Antibody eluted in this manner was greater than 96% pure as assessed by SDS-PAGE. A polyclonal sheep anti-FITC antibody was also purified from serum on the same immunosorbent to greater than 94% purity. This simple and rapid method for the purification of anti-FITC antibodies will find applications in both immunodiagnostic procedures and in studies of hapten-antibody interactions. The affinity constant of the purified monoclonal anti-FITC antibody conjugated to horseradish peroxidase was assessed by ELISA and was found to be 1.5 x 19(9) M-1.

References

Oct 1, 1989·Molecular Immunology·E W VossD W Ballard
Aug 15, 1989·Journal of Immunological Methods·I J Harmer, D Samuel
Nov 1, 1989·Journal of Virological Methods·K E BrownD Samuel
Jul 1, 1989·Journal of Bioluminescence and Chemiluminescence·D Ireland, D Samuel
Mar 16, 1988·Journal of Immunological Methods·D SamuelR A Abuknesha
Nov 1, 1986·Journal of Biochemical and Biophysical Methods·R S NiedbalaK J Schray
Jan 19, 1971·Biochemistry·D E Lopatin, E W Voss
Dec 1, 1963·The Biochemical Journal·N M GREEN

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