Patent ID: 8586294

Claim:
A method to assess intracellular translocation of a protein of interest in a mammalian cell from a first subcellular compartment to a second subcellular compartment, comprising: (a) expressing a fusion protein in said first subcellular compartment, said fusion protein comprising said protein of interest and a β-galactosidase alpha fragment which either is (i) a truncation as shown in SEQ ID NO: 6 or (ii) is less than 56 amino acids in length and comprises an amino acid sequence as shown in SEQ ID NO: 1, provided that it has a point mutation in a region spanning residues H31 and E41; (b) expressing in said cell a β-galactosidase omega fragment localized to either (i) said first subcellular compartment or (ii) said second subcellular compartment, whereby (c) translocation of said fusion protein from said first subcellular compartment to said second subcellular compartment (i) decreases local concentration of said fusion protein and said omega fragment when the omega fragment is localized to the first subcellular compartment or (ii) increases local concentration of said fusion protein and said omega fragment when the omega fragment is localized to the second subcellular compartment, whereby (d) formation of active β-galactosidase occurs when said fusion protein and said omega fragment are in the same subcellular compartment, and (e) increasing or decreasing local concentration in step (c) results in a detectible signal from active β-galactosidase proportional to an amount of translocation.