A convenient ninhydrin assay in 96-well format for amino acid-releasing enzymes using an air-stable reagent
Gerrit Haeger, Johannes Bongaerts, Petra Siegert
Abstract
An improved and convenient ninhydrin assay for aminoacylase activity measurements was developed using the commercial EZ Nin™ reagent. Alternative reagents from literature were also evaluated and compared. The addition of DMSO to the reagent enhanced the solubility of Ruhemann's purple (RP). Furthermore, we found that the use of a basic, aqueous buffer enhances stability of RP. An acidic protocol for the quantification of lysine was developed by addition of glacial acetic acid. The assay allows for parallel processing in a 96-well format with measurements microtiter plates.
Topics & Concepts
NinhydrinReagentChromatographyChemistryAcetic acidColorimetrySolubilityColorimetric analysisEnzymeAqueous solutionBiochemistryAmino acidOrganic chemistryChemical Synthesis and AnalysisAmino Acid Enzymes and MetabolismEnzyme Catalysis and Immobilization