Abstract
Fluorescence labeling of a cytokine at a specific site is required for observing cytokine-receptor interactions in living cells at the single-molecule level. Here, we demonstrated the C-terminus-specific fluorescence labeling of histidine-tagged thrombopoietin (TPO), a ligand for Mpl, with desthiobiotin-tagged fluorescent puromycin. Fluorescent TPO, purified by tandem affinity purification, stimulated the proliferation of Mpl-expressing cells. Within 10 min of its addition, fluorescent TPO was found to be diffusely distributed on the cell membranes of Mpl-expressing cells, and gradually accumulated to form fluorescent spots. This method is applicable for studying the spatial and temporal distributions of cytokines in individual living cells.
| Original language | English |
|---|---|
| Pages (from-to) | 238-242 |
| Number of pages | 5 |
| Journal | Journal of Bioscience and Bioengineering |
| Volume | 105 |
| Issue number | 3 |
| DOIs | |
| Publication status | Published - 2008 Mar 1 |
Keywords
- affinity tag
- cytokine
- desthiobiotin tag
- fluorescence labeling method
- fluorescence microscopy
- puromycin
- tandem affinity purification
ASJC Scopus subject areas
- Biotechnology
- Bioengineering
- Applied Microbiology and Biotechnology