A self-immolative reporter for β-galactosidase sensing

A dual fluorogenic and chromogenic probe, Gal-2SBPO, has been developed based on a synthesized water-soluble dye, 9-di-3-sulfonyt-propylaminobenzo[a]phenoxazonium perchloride (2SPBO, lambda(ex/em) = 630/670 nm). beta-D-Golactopyronoside, the substrate of beta-galactosidase, was conjugated to 2SBPO through a para-substituted benzyloxycorbonyl group and a glycine residue, which serve as a self-immolative spacer and as a molecular blocker to mask the optical signal of 2SBPO, respectively. Gal-25BPO was soluble and stable under physiological conditions. Enzymatic cleavage of the P-D-galactopyranoside triggered a series of spontaneous reactions that resulted in a release of optically active 2SBPO. The beta-galactosidase activity was assayed by monitoring the absorbance at 630 nm and fluorescence at 670 nm.