Atto-643. DC-SIGN forms stable homotetramers on cell surface, mediated by the neck region coiled-coil formation. We and others have demonstrated previously that DC-SIGN extracellular domain faithfully maintains the tetramer structure and MLGI properties of the full-length protein. Hence, DC-SIGN extracellular segment (named as DC-SIGN hereafter) was used to study its MLGI properties with Gx-psDiMan in solution. To facilitate sensitive fluorescence based binding detection, the recombinant DC-SIGN mutant DC-SIGN-Q274C was expressed in E. coli and purified using Sepharose-Mannose affinity chromatography as described previously. The purified proteins were site specifically labelled with a maleimide modified Atto-643 dye (named as labelled DC-SIGN) through the Michael addition between thiol and maleimide as described previously. This labelling site is close to, but not sits in the CRD's glycan binding pocket, and thus dye-labelling does not affect CRD's glycan binding properties as confirmed previously. Atto-643 was chosen here due to its high fluorescence quantum yield, excellent photostability, and strong hydrophilicity, thereby minimizing any potential interfere with the CRD structure and glycan binding properties. Moreover, its fluorescence emission peaks at the far-red region of the visible spectrum (e.g., λEX = 630 nm, λEM ~660 nm), which can minimize the potential interference with fluorescence readout arising from the GNP's inner filter effect. The success of protein production and Atto-643 labelling were confirmed from their respective high resolution mass spectra (HRMS), where an increase of molecular mass of 935 was observed.