A new method of iodinating ovalbumin, a protein which lacks accessible tyrosine groups, by conjugation to a highly fluorescent coumarin active ester, CASE.
A simple and efficient method is described to overcome the difficulty in radiolabelling ovalbumin and is applicable to any protein. Coumarin-3-acetic acid-N-OH-succinimidyl ester (CASE) binds directly to ovalbumin in aqueous solution to form covalent bonds with free amino groups. The attached CASE residues can then be radiolabelled by the chloramine-T method. This offers an inexpensive alternative to the Bolton and Hunter reagent with the advantage that CASE is fluorescent. This property, apart from direct use as a fluorescent tag, allows one to monitor the conjugate before radiolabelling. Below a conjugation ratio of 9:1, CASE-ovalbumin retained full antigenic identity with ovalbumin.
D000818:Animals; D000937:Antigen-Antibody Reactions; D002700:Chloramines; D005456:Fluorescent Dyes; D007457:Iodine Radioisotopes; D051379:Mice; D010047:Ovalbumin / Q000276:immunology; D014105:Tosyl Compounds; D014468:Umbelliferones
D002700:Chloramines; D005456:Fluorescent Dyes; D007457:Iodine Radioisotopes; D014105:Tosyl Compounds; D014468:Umbelliferones; C016300:chloramine-T; D010047:Ovalbumin; C044852:7-hydroxycoumarin-3-acetic acid N-hydroxysuccinimidyl ester