Use of lectins for probing differentiated human embryonic stem cells for carbohydrates
2006
Wearne, Kimberly A. | Winter, Harry C. | O'Shea, Katherine | Goldstein, Irwin J.
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 α-D-mannopyranosyl groups, poly-N-acetyllactosaminyl chains, both α2,3- and α2,6-linked N-acetylneuraminic acid (Neu5Ac), α1,6-linked L-fucosyl, and β-D-galactosyl groups, and more specifically, the T, Tn, and sialyl-Tn antigens. However, no α1,2-linked L-fucosyl, terminal nonreducing α-D-galactosyl, N-acetyl-β-D-glucosaminyl, nor N-acetyl-α-D-galactosaminyl groups were found by this approach. We also established the presence of Neu5Acα2,3/2,6-Galβ1,4 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 α2,3-linked Neu5Ac as well as a change to exclusively α1,6-linked L-fucose upon differentiation.
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