"
Team:British Columbia/Notebook/Protocols/BiopanningAssay
From 2014.igem.org
Biopanning Assay
Unfixed chalcopyrite:
- Pre-sieve chalcopyrite with 40um size cell strainer. Then autoclave by Dry-5 times settings
- Incubate chalcopyrite (40um in size, 30mg) in culture of Caulobacter crescentus for 2hrs, shaking at low rpm.
- Filter chalcopyrite + Caulobacter crescentus using a 40um Falcon™ Cell Strainers
- Washing steps:
- 1st round of panning: Wash strained chalcopyrite with 5ml 1x Tris-buffered saline (TBS) containing 0.1% (v/v) Tween-20, pH 7.2. Plate cells.
- 2nd round of panning: Wash strained chalcopyrite with 5ml 1x Tris-buffered saline (TBS) containing 0.2% (v/v) Tween-20, pH 7.2.
- 3rd round of panning: Wash strained chalcopyrite with 5ml 1x Tris-buffered saline (TBS) containing 0.3% (v/v) Tween-20, pH7.2.
- 4th round panning: Wash strained chalcopyrite with 5ml 1x Tris-buffered saline (TBS) containing 0.4% (v/v) Tween-20, pH7.2.
- 5th round panning: Wash strained chalcopyrite with 5ml 1x Tris-buffered saline (TBS) containing 0.5% (v/v) Tween-20, pH7.2
