Fig. 1.

Detecting protein functionalization by loss of free amines via the colorimetric ninhydrin assay. (a) Reaction scheme for chemical functionalization of a protein by anhydrides or succinimidyl esters, which occurs primarily at free amines. (b) Structures of reaction products of amine sidechains with methacryloyl or norbornene groups. (c) Reaction scheme for the ninhydrin assay. Ninhydrin reacts with free primary amines to generate Ruhemann’s Purple, a purple-colored soluble product. (d) Photo of well-plate after running the optimized assay with a gelatin standard curve. Solutions remained clear, free of precipitation, yielded a visible color change that corresponded to free amine content in the solution. (e) Photo of well plate after analyzing functionalized gelatin samples (here, GelMA). Samples yielded color changes that inversely correlate with their respective DoF values.