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HAI-2 species in Caco-2 human enterocytes.

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posted on 2017-01-26, 00:39 authored by Frank Shiao, Li-Ching O. Liu, Nanxi Huang, Ying-Jung J. Lai, Robert J. Barndt, Chun-Che Tseng, Jehng-Kang Wang, Bailing Jia, Michael D. Johnson, Chen-Yong Lin

Caco-2 human enterocytes were incubated in PBS as a non-activation control (lanes 1) or a phosphate buffer pH 6.0 for 20 min to induce matriptase zymogen activation (lanes 2). Matriptase species were subsequently immunodepleted from the lysates that had been prepared from Caco-2 cells in which matriptase zymogen activation was induced (lanes 3). The three Caco-2 lysates were then analyzed by immunoblot for HAI-2 species with significant N-glycan branching using HAI-2 mAb DC16 (HAI-2 DC16), HAI-2 species without N-glycan branching using HAI-2 mAb XY9 (HAI-2 XY9), and matriptase species using matriptase mAb M24 (Total MTP). Small quantities of mouse IgG shed from the beads used for immunodepletion can be seen as a band of approximately 150-kDa (lanes 3 in each panel) is indicated by an arrow. The HAI-2 species in Caco-2 cells have been observed in at least 10 independent experiments. The induction of matriptase zymogen activation in Caco-2 cells and the immunodepletion of matriptase species has been carried out at least 3 times. Representative data are shown.