title: New tetrazolium method for phosphatase assay using ascorbic acid 2-phosphate as a substrate.
abstract: A new method to assay alkaline and acid phosphatases assay using ascorbic acid 2-phosphate (AsA-P) and ditetrazolium salt nitroblue tetrazolium chloride (NBT) was developed. AsA-P is hydrolyzed in the presence of phosphatase to yield ascorbic acid. In turn, the ascorbic acid reduces NBT directly or indirectly, opening the tetrazole ring to produce an insoluble formazan as a colored precipitate. The proposed method for alkaline phosphatase was compared with a conventional method in which 5-bromo-4-chloro-3-indolyl phosphate (BCIP) is used in combination with NBT in the dot blots of a dilution series of beta-lactoglobulin. AsA-P reduced NBT more effectively than BCIP in the presence of alkaline phosphatase. AsA-P could be also used as the chromogenic substrate for an acid phosphatase assay in the presence of phenazinium methylsulfate and NBT.
authors.affiliation: Biotechnology and Food Research Institute, Fukuoka Industrial Technology Center, Kurume 839-0861, Japan.
authors.forename: Tadayuki
authors.initials: T
authors.lastname: Tsukatani
authors.forename: Seiji
authors.initials: S
authors.lastname: Ide
authors.forename: Masashi
authors.initials: M
authors.lastname: Ono
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Matsumoto