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| Shin, In-Sun; Nishikawa, Kiyotaka; Maruyama, Hiroki; Ishii, Satoshi. |
| A two-step binding assay for globotriaosylceramide (Gb3) content was developed by histidine-tagging strategy, which is a well-established method for the purification of recombinant proteins. The complete binding of the recombinant His-tagged Shiga toxin 1B subunit (1B-His) (1 microg/ml) to the standard Gb3 adsorbed on a multi-well H type plate was observed within 30 min at 37 degrees C; and its binding could be visualized by the following applications of HisProbe-HRP (8 microg/ml) and tetramethylbenzidine (TMB) peroxidase substrate. The 1B-His binding assay was linear over the range of 1 to 100 ng of Gb3 per well. The binding of 1B-His was specific to Gb3 separated from HeLa cells, and no major cross-reactivity of other glycolipids in Folch's lower... |
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Palavras-chave: Globotriaosylceramide; Shiga toxin; Binding assay method; His-tagged protein. |
| Ano: 2006 |
URL: http://ir.obihiro.ac.jp/dspace/handle/10322/833 |
