Possible causes of error

Possible causes of error

1. Size-Exclusion Chromatography

   • column wasn't completely vertical
   • column bed wasn't completely flat
   • column too short to allow protein to run fully

2. Ion-exchange chromatography

   • only collected two fractions
     • couldn't create a curve for ion chromatography
   • not collected fast enough after adding high salt

3. incorrect buffer solution denatured protein

   • buffer of pH not exact: 8.95 rather than 8.9
     • Pasteur pipette added too high volume of NaOH - needed finer control of volume addded
   • inaccuracies of pH metre

4. ultracentrifugation of sample

   • membrane of centrifugal concentrator damaged
     • HRP leaked through membrane
   • too much centrifugation denatured protein