Detergents are often used to solubilize membrane proteins prior to biochemical investigation. To measure binding affinities using temperature-related intensity change (TRIC) or Spectral Shift, the first step is to label one of the binding partners. If this molecule is a membrane protein solubilized in detergent at a concentration above the CMC, it may be necessary to adapt the labeling protocol. Free detergent micelles can capture excess free dye molecules resulting in 1) a lower dye concentration available in the labeling reaction and 2) the presence of micelle-captured free dye during the binding affinity measurement. This can lead to decreased labeling efficiency and signal quality.  This protocol was tested for NHS, Maleimide and tris-NTA labeling, but can be adjusted for other labeling strategies. A more general description can be found in Labeling Membrane Proteins for Binding Assays.

Membrane proteins | Labeling | Purification | Size Exclusion Chromatography | MOX-P-106