User:Samiye Yaman/Notebook/Lab 571/2011/09/28

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Entry title

Objective

Description

- Two buffer solution (one with 50 mL of Tris and one with 50mL of Tris plus 1M of NaCl)was filtered. The one without NaCl was filtered first.


- Dialysis tubes were taken out of the buckets and the solution inside of the dialysis tubes were poured in to a beaker.


-Fast protein liquid chromatography was used.

   *Buffer A(50 mM of Tris) was loaded to the loop.
   *Protein was loaded to the loop.
   *Buffer B (50 mM of Tris+ 1M Nacl) was loaded to the loop.
     **Both Buffer A and B were covered with aluminum foil to prevent the entry of foreign particles.
   *Buffer B was run in order to get rid of the foreign particles inside the column.
   *Buffer A was run again to start the procedure.
   *10 ml The loop was injected to was the negatively charged particles in the column.
   *The gradient was run at 50% for 15min.
   *Fractions (5ml each) were collected.
   *The gradient was continued to run for 50% for 15 more mins again.
   *Fractions (5ml each)were collected.
   *The gradient was run at 50% for 25min.
   *Fractions (5ml each) were collected.
   *Rest of the protein was loaded to the loop.
   *The gradient was run with 100% in order to clean the column.
   *The fraction was run at 0.
   *Buffer B was run again.
   *After running Buffer B, Buffer A was run in order to reequilibrate.


-Fractions belongs to peak 1, peak 2 and peak 3 were collected and placed in to 3 separate test tubes.

Image:Untitled.jpg

Data

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Notes

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