Two years ago, Bertozzi and her group used their copper-free click chemistry technique to image cells in live zebrafish embryos, which by virtue of being transparent, are popular for scientists studying embryogenesis. Those studies revealed dramatic differences in cell-surface expression, intracellular trafficking patterns, and tissue distributions of glycans at different stages of zebrafish larval development.
"However, we were unable to detect labeled glycans in zebrafish embryos earlier than 24 hours post-fertilization," Bertozzi says. "Because many important developmental events including cell migration, tissue morphogenesis, and cell differentiation occur in the first 24 hours of zebrafish embryogenesis, and glycan biosynthesis is also known to occur within that 24 hour period, we sought to develop methods that would enable us to image the glycans early during embryogenesis."
For this study, Bertozzi and her co-authors microinjected embryos with azido sugars at the one-cell stage, allowed the zebrafish to develop, and then detected the metabolically labeled glycans with the copper-free click chemistry technique. This enabled them to image certain types of glycans as early as seven hours post-fertilization. In addition, they used a complementary, non-metabolic labeling technique to target a class of glycans that carry sialic acid, giving them simultaneous but independent ways to image two distinct classes of glycans.
Time-lapse and multicolor imaging experiments highlighted differences between the O-glycans and sialylated glycans in the cells during the gastrulation and segmentation periods of embryogenesis. The results revealed a dramatic re-or
|Contact: Lynn Yarris|
DOE/Lawrence Berkeley National Laboratory