Freimoser:Protocols:ProteinQuantification: Difference between revisions

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==Absorbance assays==
==Absorbance assays==


Revision as of 08:04, 30 October 2006

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Absorbance assays

Absorbance at 280 nm

  • Range: 20 micrograms to 3 mg
  • Volume: Depends on cuvette - volumes range from 200 microliters to 3 ml or greater
  • Accuracy: Fair
  • Convenience: Excellent, if equipment available
  • Major interfering agents: Detergents, nucleic acids, particulates, lipid droplets

Absorbance at 205 nm

  • Range: Roughly 1 to 100 micrograms
  • Volume: Depends on cuvette - volumes range from 200 microliters to 3 ml or greater
  • Accuracy: Fair
  • Convenience: Very good
  • Major interfering agents: Detergents, nucleic acids, particulates, lipid droplets

Extinction Coefficient

  • Range: 20 micrograms to 3 mg
  • Volume: Depends on cuvette - volumes range from 200 microliters to 3 ml or greater
  • Accuracy: ~2% (very good)
  • Convenience: Very good
  • Major interfering agents: Detergents, nucleic acids, particulates, lipid droplets

Colorimetric assays

Modified Lowry

  • Range: 2 to 100 micrograms
  • Volume: 1 ml (scale up for larger cuvettes)
  • Accuracy: Good
  • Convenience: Fair
  • Major interfering agents: Strong acids, ammonium sulfate

Biuret

  • Range: 1 to 10 mg
  • Volume: 5 ml (scale down for smaller cuvettes)
  • Accuracy: Good
  • Convenience: Good
  • Major interfering agents: Ammonium salts

Bradford assay

  • Range: 1 to 20 micrograms (micro assay); 20 to 200micrograms (macro assay)
  • Volume: 1 ml (micro); 5.5 ml (macro)
  • Accuracy: Good
  • Convenience: Excellent
  • Major interfering agents: None

Bicinchoninic Acid (Smith)

  • Range: 0.2 to 50 micrograms
  • Volume: 1 ml (scale up for larger cuvettes)
  • Accuracy: Good
  • Convenience: Good
  • Major interfering agents: Strong acids, ammonium sulfate, lipids

Amido Black method

  • Range: 2 to 24 micrograms
  • Volume: 2 ml
  • Accuracy: Good
  • Convenience: Poor
  • Major interfering agents: None reported

Colloidal Gold

  • Range: 20 to 640 nanograms
  • Volume: 1 ml (scale up for larger cuvettes)
  • Accuracy: Fair
  • Convenience: Poor
  • Major interfering agents: Strong bases
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