NOVEL METHOD TO DETECT BETA-GALACTOSIDASE BY A DOT-BLOTTING ASSAY ON NITROCELLULOSE MEMBRANE USING 6-BROMO-2-NAPHTHYL BETA-D-GALACTOPYRANOSIDE AS SUBSTRATE

A novel method to detect beta-galactosidase by dot-blotting samples on nitrocellulose membrane and staining the membrane with azo dye coupli ng technique using 6-bromo-2-naphthyl beta-D-galactopyranoside as subs trate is presented. The enzymatic reaction to split the substrate is p H-dependent, and the sensitivity of this method could be increased by more than 10-fold with an additional treatment of 0.01 M sodium carbon ate after the coupling of diazo-blue B with 6-bromo-2-naphthyl release d by beta-galactosidase. As low as 0.0015 unit of beta-galactosidase c ould be detected within 10 min. This improved beta-galactosidase stain ing method may be used for a native or IEF gel to show the beta-galact osidase active band and may also detect other hydrolases using substra tes for the azo dye coupling technique.