	US 12,227,787 B2
	Method of evaluating quality of dephosphorylation reagent and method of detecting target nucleic acid
Masateru Ito, Kamakura (JP); Yoji Ueda, Kamakura (JP); Yuki Takii, Kamakura (JP); and Mai Yagi, Kamakura (JP)
Assigned to Toray Industries, Inc., Tokyo (JP)
Appl. No. 17/280,056
Filed by Toray Industries, Inc., Tokyo-to (JP)
PCT Filed Sep. 25, 2019, PCT No. PCT/JP2019/037518 § 371(c)(1), (2) Date Mar. 25, 2021, PCT Pub. No. WO2020/067120, PCT Pub. Date Apr. 2, 2020.
Claims priority of application No. 2018-179529 (JP), filed on Sep. 25, 2018.
Prior Publication US 2022/0033874 A1, Feb. 3, 2022
This patent is subject to a terminal disclaimer.
Int. Cl. C12Q 1/42 (2006.01); C12N 9/16 (2006.01); C12Q 1/6816 (2018.01); G01N 30/72 (2006.01); G01N 30/74 (2006.01); G01N 30/86 (2006.01); G01N 33/68 (2006.01); G01N 30/02 (2006.01)

CPC C12Q 1/42 (2013.01) [C12N 9/16 (2013.01); C12Q 1/6816 (2013.01); G01N 30/7233 (2013.01); G01N 30/74 (2013.01); G01N 30/8679 (2013.01); G01N 33/6848 (2013.01); C12Y 301/03001 (2013.01); G01N 2030/027 (2013.01); G01N 2410/00 (2013.01)]

7 Claims

1. A method of evaluating a quality of a dephosphorylation reagent, the method comprising the steps of:

making a dephosphorylation reagent comprising an alkaline phosphatase and a peptide fragment group (D1) composed of two or more peptide fragments, wherein each of the two or more peptide fragments consists of 5 to 50 consecutive amino acid residues selected from positions 501 to 578 of the amino acid sequence set forth in SEQ ID NO: 10;

determining a content ratio of the peptide fragment group (D1) to the alkaline phosphatase using formula (H1): (X_D1/Y)×100<4.4000 (H1),

wherein X_D1represents a peak area value of the peptide fragment group (D1) calculated by an automatic integration method from an extracted ion chromatogram obtained by an LC-MS/MS analysis of the dephosphorylation reagent, and Y represents a peak area value of the alkaline phosphatase calculated by an automatic integration method from a chromatogram obtained by an LC-UV analysis of the dephosphorylation reagent; and

evaluating the dephosphorylation reagent as having a high quality if the content ratio of the peptide fragment group (D1) to the alkaline phosphatase satisfies formula (H1).