Lissamine Rhodamine Research Articles

Purification of Fluorescent Protein Conjugates : Comparison of Charcoal and ‘Sephadex’

REMOVAL of unreacted fluorescent material from fluorescent protein conjugates is essential for unambiguous interpretation of microscopical findings in immunofluorescence1. Many procedures have been suggested for conjugate purification, but extraction with powdered activated charcoal2 and gel filtration with ‘Sephadex’3,4 are at present the only efficient methods suitable for general application. These two methods have been compared as purification procedures for conjugates of lissamine rhodamine B (RB 200) and fluorescein isothiocyanate. We have confirmed the inefficiency of dialysis5: after exhaustive dialysis (7–10 days), conjugates still contain appreciable unreacted fluorescent material which can be removed by shaking with charcoal or passage through ‘Sephadex’.