A glycated albumin (GA) in a blood cell reflects a history (of about two or three weeks) of an in-vivo blood glucose level, and thus is used as an important indicator for diagnoses, treatment and the like of diabetes. A glycated albumin is an albumin whose an ε-amino group of a lysine residue is glycated, and an amount (%) thereof is represented by a ratio (%) of the glycated albumin amount with respect to a total albumin amount.
As a method for assaying the glycated albumin, high performance liquid chromatography (HPLC) and the like are exemplified generally, but as a simple and low-cost method, the following enzymatic method is utilized practically. In this method, firstly, a fructosyl amino acid oxidase (hereinafter, referred to as a “FAOD”) is allowed to act on a glycated part of the albumin, thereby generating hydrogen peroxide. An amount of this hydrogen peroxide corresponds to a glycation amount of the albumin. Further, a peroxidase (hereinafter, referred to as a “POD”) and a chromogenic substrate that develops a color by oxidation are added to this reaction liquid, so as to effect a redox reaction between the hydrogen peroxide and the chromogenic substrate with the POD as a catalyst. Then, the glycation amount of the albumin is obtained by measuring a chromogenic level of the chromogenic substrate, so that a ratio of the glycated albumin (hereinafter, also referred to as a “GA (%)”) can be calculated from the glycation amount and the total albumin amount.
As described above, since the glycated albumin is characterized by the glycation of the ε-amino group of the lysine residue, it is desired that the FAOD acts on the glycated lysine residue efficiently. However, this FAOD is not likely to act directly on a protein, and generally, a digestion product of the albumin is prepared by protease treatment so that the FAOD is allowed to act thereon.
However, the albumin in a blood sample is different from an albumin as a purified reagent in that various substances, for example, bilirubin, a lipoprotein and the like bind thereto, so that it is difficult to obtain a fragment on which the FAOD is likely to act from protease treatment. Thus, when assaying an albumin in a sample such as a serum and a blood plasma, it is necessary to add a large amount of the protease into the sample so as to raise reactivity (Patent Document 1), but this causes problems as listed below.
An enzyme and the like in other reagent used for assaying the glycated albumin are deactivated by the presence of the large amount of the protease.
In the case where the large amount of the protease is contained in a reagent, when only a slight part of the reagent contaminates other assaying reagent, an enzyme and an antibody in the other assaying reagent are deactivated, and the other assaying reagent cannot be used.
A protease solution with a high concentration is necessary for adding the large amount of the protease, but the protease is autolyzed so as to degrade the stability.
In the case of seeking to assay in a dry system, for example, preparation of a protease solution with a high concentration that is necessary for preparing a specimen is difficult.
High cost is required for using the large amount of the protease.                Patent document 1: WO 2002/061119 pamphlet        
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