IGEM:Harvard/2006/Adaptamers/Notebook/2006-8-26

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Revision as of 18:33, 26 August 2006

8/26

Strand Displacement

We'll test if strand displacement can quench the adaptamers.

  • Abbreviations/Syntax

SMB: Streptavidin Magnetic Beads BB: Bittker Buffer SS: standard setup r A B: replace A in SS with B.

  • Conditions

1. Standard setup: 40 pmol A20 + 100uL .4% solids SMB + 160 pmol thrombin 2. =1 3. +10 pmol A20.20 4. + 20 pmol A20.20 5. + 40 pmol A20.20 6. + 10 pmol A20.35 7. + 20 pmol A20.35 8. + 40 pmol A20.35 9. r A20 A50 10. r A20 T20 11. r A20 S20 12. r thrombin nothing

  • Methods

100 uL SMB --> 10 uL + 40pmol (1-8, 12. A20 9. A50 10. T20 11. S20 12. A20) + (1-11. 160 pmol thrombin) --> 15 uL -->30 minute shake --> + (3. 10 pmol A20.20 4. 20 pmol A20.20 5. 40 pmol A20.20 6. 10 pmol A20.35 7. 20 pmol A20.35 8. 40 pmol A20.35) --> 19 uL --> shake 20 minutes --> 2X wash --> + 4 uL .200 ug/uL anti-thrombin primary Ab --> 2X wash --> + 1 ug/uL seconday Ab --> 3X wash --> 50 uL --> imaging on 96 well plate.

  • Results


ELISA

We'll attempt to quantify how much thrombin is bound by the adaptamers using an ELISA

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