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Esults. Embryos at high stage (3 1/3 hpf), 50 epiboly (5 1/4 hpf), 70 epiboly (7 hpf) and tailbud stage (10 hpf) were deyolked, separated by SDS-gel electrophoresis and Coomassie stained (A) or blotted and immunodetected with antibodies against Tubulin (55 kD) and Moesin (78/80 kD apparent molecular weight) (B). Note that total protein amount was lower in deyolked samples, therefore more embryos
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Of each cell colony over 24 hours was calculated. Data for each cell line was averaged over 10 wells and compared between parental, control transfected, and galectin-1 transfected cells. A t-test was applied to compare means.In vitro invasion assay6.8 pg +/- 4.2 pg. In spite of this variability, the quality of the RNA was consistently high with a mean RNA integrity number of 8.13 ?0.74. Expression
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Horesis The establishment of the deyolking protocol was a prerequisite for high quality 2D gels from early zebrafish embryos. After removal of the predominant yolk proteins we were able to generate high resolution 2D gels in the acidic (pI 4?, Fig. 4A) as well as in the basic range (pI 6?9, Fig. 4B). We established a protocol that is compatible with three colour fluorescent labelling using the Ett
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By the ample amount of normal mouse brain tissue available for dissection. In spite of species differences, cross-hybridization of mouse genetic material to human probes did prove to be a common occurrence. These data made it possible to control, rather stringently, for the potential contamination of tumor edge samples with mouse brain. Of course, there could still be possible contamination ?react
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E study targeted larval stages 48 or 72 hpf (hours post fertilization), when the yolk to cell mass ratio is already decreased [5], however, without identifying the proteins. Therefore, it remains unclear whether at this stage analysis without deyolking provides satisfactory information about cellular proteins. Thus, the develop-ment of a reliable method to remove the interfering yolk from cells on
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Of each cell colony over 24 hours was calculated. Data for each cell line was averaged over 10 wells and compared between parental, control transfected, and galectin-1 transfected cells. A t-test was applied to compare means.In vitro invasion assay6.8 pg +/- 4.2 pg. In spite of this variability, the quality of the RNA was consistently high with a mean RNA integrity number of 8.13 ?0.74. Expression
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Yolk removal. Embryos with yolk (Y) were analysed in comparison with embryos after one-step deyolking (D) or after two additional wash steps (W). A. Total protein amount per embryo as determined by DC protein assay (Bio-Rad). B. Coomassie stain (0.5 embryos loaded per lane). C. Western blot against Tubulin and MEK (0.5 embryos loaded per lane). While yolk proteins were efficiently depleted, recove
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Ed clones were compared to their GFP control counterparts. (Westerns controlled for loading by -actin IB). (D) Over-expression of galectin-1 promotes invasion. All cell counts were normalized to the parental cell line data. (Westerns controlled for loading by -actin IB).our identification of galectin-1 as a mediator of glioma invasion has been corroborated previously as detailed below. While previ

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