Journal of Experimental Medicine, Vol 145, 443-449, Copyright © 1977 by Rockefeller University Press
Isolation by cell-column chromatography of immunoglobulins specific for cell surface carbohydrates
BA Sela and GM Edelman
A new method of affinity chromatography using glutaraldehyde-fixed cells
immobilized on Sephadex beads has been used to isolate immunoglobulins
(Ig's) specific for cell surface glycoproteins. Ig's that specifically
bound and agglutinated the same cells as those originally fixed on the
columns were isolated from nonimmune sera of various species. Periodate
treatment of the cell-columns and the free cells destroyed their ability to
bind the Ig's, and the binding of the Ig's to untreated cells was inhibited
by monosaccharides such as D- galactose and sialic acid. The binding of
antibodies directed against cell surfaces obtained by immunizing animals
with the same mouse tumor cell lines used on the columns (P388 and EL4) was
not inhibited by various saccharides. Surface glycoproteins obtained from
the mouse tumor cells by immunoprecipitation with the column-isolated Ig's
yielded specific electrophoretic patterns that differed from those obtained
using Ig's from the sera of rabbits immunized with the tumor cells. The
data suggest that the Ig's isolated by cell-column chromatography were
directed against carbohydrates, probably those in terminal positions of the
polysaccharide portions of the tumor cell surface glycoproteins.
Column-isolated Ig's specific for carbohydrates were also useful in studies
of cell interactions in nonmammalian systems including Dictyostelium
discoideum and Saccharomyces cerevisiae. The cell-column method appears to
be adaptable to the isolation of a variety of molecules in addition to
antibodies.
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