A340 nm assays of ACS and ADH

07 May 2015

Amount of protein to use

• ACS assays:
  • 5/5/2015:
• ADH assays:
• In SIP:
  • 2015/5/13: 45uM ACS worked just enough to detect ADH activity over 30 min.

Amount of NADH to use

• In SIP, ~4mM seems appropriate.
  • SIP extracts include FDH and do not run out of NADH.
    • See: /Users/jmatsen/Dropbox/In_Progress/CO2_goes_to_methylotrophy/Enzyme_Assays/Nash_SIP3-4/2015_05_13 SIP3-4 ADH assay

Amount of cofactors to use

• Reasonable as of 5/14/2015
  • 1 mM CoA, 4 mM ATP, 5mM MgCl2

Tips

• Use of UV-transparent plastic enhances the range at which you can monitor over
  • Polystyrene has background absorbance of ~0.17 A340 units on our machine. (source data)
  • UV-star plates have a background absorbance of __ on our machine.

Other resources

• wiki: :
  • Molecular Devices Plate Reader
    • Shows
• Old records:
  • 2015 Nash ACS assay screening records summary