Figure 2
From: Spatial separation and bidirectional trafficking of proteins using a multi-functional reporter
Targeting HaloTag protein to the cell surface. (a) Human Ī²1 integrin is depicted with a signal peptide (SP), extracellular domain (EC), transmembrane domain (TM) and cytoplasmic domain (CD) (left). In the Ī²1Int-HaloTag construct, the HaloTag protein is displayed on the cell surface by fusion to the truncated (tr) extracellular domain of human Ī²1 integrin (right). (b) Immunocytochemistry, without permeabilization, using antibodies against integrin (green) and HaloTag (red) show that the Ī²1Int-HaloTag fusion protein is being expressed in transfected HeLa Hcells, and the overlap of the two proteins (yellow) indicates Ī²1Int-HaloTag is expressed on the cell surface in a similar pattern to endogenous Ī²1 integrin. (c) HEK293 cells stably expressing Ī²1Int-HaloTag labeled with the HaloTag 488 ligand show a green rim, confirming that HaloTag is on the surface and functional. (d) HEK293 cells stably expressing HaloTag-(NLS)3 labeled with the HaloTag 488 ligand do not show a green rim, confirming that the ligand is cell impermeable and does not label intracellular HaloTag protein. Cell images were generated on an Olympus FV500 confocal microscope using the appropriate filter.