title: Use of lectins for probing differentiated human embryonic stem cells for carbohydrates.
abstract: The carbohydrates present on the surface of differentiated human embryonic stem cells (hESCs) are not yet well established. Here, we have employed a panel of lectins and several anti-carbohydrate antibodies to determine the carbohydrates that are present at day 12 of hESC differentiation as embryoid bodies (EBs). On the basis of staining with fluorescein-labeled lectins, we have determined the presence of both terminal and internally linked alpha-d-mannopyranosyl groups, poly-N-acetyllactosaminyl chains, both alpha2,3- and alpha2,6-linked N-acetylneuraminic acid (Neu5Ac), alpha1,6-linked l-fucosyl, and beta-D-galactosyl groups, and more specifically, the T, Tn, and sialyl-Tn antigens. However, no alpha1,2-linked l-fucosyl, terminal nonreducing alpha-D-galactosyl, N-acetyl-beta-D-glucosaminyl, nor N-acetyl-alpha-D-galactosaminyl groups were found by this approach. We also established the presence of Neu5Acalpha2,3/2,6-Galbeta1,4 GlcNAc-terminated chains on the surfaces of 12-day-old EBs, as indicated by the great enhancement of staining by Erythrina cristagalli agglutinin (ECA) after treatment with neuraminidase. In each case, inhibition of binding by a haptenic sugar or treatment with neuraminidase was used to eliminate the possibility of nonspecific binding of the lectins. A comparison with undifferentiated cell staining revealed an increase in alpha2,3-linked Neu5Ac as well as a change to exclusively alpha1,6-linked l-fucose upon differentiation.
authors.affiliation: Department of Biological Chemistry, University of Michigan, Ann Arbor, MI 48109, USA.
authors.forename: Kimberly A
authors.initials: KA
authors.lastname: Wearne
authors.forename: Harry C
authors.initials: HC
authors.lastname: Winter
authors.forename: Katherine
authors.initials: K
authors.lastname: O'Shea
authors.forename: Irwin J
authors.initials: IJ
authors.lastname: Goldstein
mesh_terms: D002241:Carbohydrates / Q000032:analysis*; D002454:Cell Differentiation; D004622:Embryo, Mammalian / Q000166:cytology; D004627:Embryonic Induction; D053595:Embryonic Stem Cells / Q000737:chemistry / Q000166:cytology*; D005455:Fluorescent Antibody Technique / Q000379:methods; D006801:Humans; D037121:Plant Lectins / Q000494:pharmacology*
chemical_list: D002241:Carbohydrates; D037121:Plant Lectins