JP2003259896A - Aspartic acid aminotransferase activity-measuring reagent - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide an aspartic acid aminotransferase activity-measuring reagent which can inhibit the increase of a reagent blank reaction, namely the increase of an initial absorbance, and to provide a method for measuring the same. <P>SOLUTION: This aspartic acid aminotransferase activity-measuring reagent comprising L-aspartic acid, 2-oxoglutamic acid, a malic acid dehydrogenase, a lactic acid dehydrogenase, and a reducing type nicotinamide adenine dinucleotide is characterized by further containing a substance having an action for inhibiting the activity of the lactic acid dehydrogenase. <P>COPYRIGHT: (C)2003,JPO

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a reagent for measuring aspartate aminotransferase activity.

[0002]

2. Description of the Related Art Aspartate aminotransferase (hereinafter abbreviated as AST) is an enzyme that is widely distributed in the heart and liver and is translocated into the blood during various diseases.
The measurement of AST activity in biological fluids such as urine and blood is one of the important items as an index for the follow-up observation of diagnosis or treatment of heart disease or liver disease.

A method for measuring AST activity is to use AS-aspartic acid and 2-oxoglutarate as substrates.
Oxaloacetate produced by T is converted to malate by malate dehydrogenase (hereinafter abbreviated as MD),
The reduced amount of reduced nicotinamide adenine dinucleotide (hereinafter, abbreviated as NADH) that was allowed to coexist was
A method of measuring AST activity by measuring at a wavelength near 340 nm is widely used.

The reaction equation is shown below. In the above formula, NAD is oxidized nicotinamide adenine dinucleotide.

In the AST activity measuring method based on the above reaction formula, when a test sample (for example, blood) containing pyruvate and lactate dehydrogenase (hereinafter abbreviated as LD) is measured, NADH in the reagent is Since the AST activity decreases independently of the AST activity, a positive error is given to the AST activity measurement. In order to avoid this phenomenon, a method is used in which a large amount of LD is added together with NADH and pyruvate derived from the test sample is consumed before the AST reaction is started to avoid the effect. The reaction formula is as follows.

[0006] As described above, the AST activity measurement is one of the important items as an index for diagnosing heart disease or liver disease and observing the course of treatment, and is therefore measured internationally.
However, since various measuring methods of AST activity are known and various measuring methods are used, the measured values are different between facilities and measurers, and the compatibility of the measured values is difficult.
May impair clinical diagnosis. Therefore, in each country, the recommended method that stipulates the reaction principle, reagent composition, reagent concentration, etc. has been proposed, and AST activity measurement is performed at each facility to obtain compatibility with the measured value obtained by the recommended method. I'm starting to be seen.

In Japan, the Japanese Society for Clinical Chemistry (JSCC)
In 1989, the recommended method for measuring AST activity was published [The Japan Society for Clinical Chemistry: Recommended Method for Measuring Enzyme Activity in Human Serum-Aspartate Aminotransferase- (1989-0.
8-30), clinical chemistry, 18 (4), 226-249.
(1989)]. The recommended method published by the Japan Society for Clinical Chemistry is a method for measuring common enzyme activity under optimal conditions that can be shared at the technical level of clinical laboratories, etc., in order to ensure compatibility of measured values. Therefore, it cannot be generally used as a daily inspection method. Therefore, clinical test drug manufacturers provide reagents that can be compatible with the recommended method of the Japanese Society for Clinical Chemistry and measured values.
And, on the assumption that the recommended method of the Japan Society for Clinical Chemistry and the measured value are compatible, we make an effort every day to provide a more stable and inexpensive reagent that can obtain an accurate and precise measured value. ing.

[0008]

As described above, in a facility that measures AST activity such as a clinical laboratory, it is possible to obtain compatibility between the recommended method of the Japan Society for Clinical Chemistry and the measured value as a daily test method. It is common to use an AST activity measuring reagent provided by a clinical test drug manufacturer. However, biological samples are a mixture of many different components, and
Since the AST activity measuring reagent is also a mixture of various components, it is extremely difficult to develop a perfect AST activity measuring reagent.

Particularly, in an analyzer called an automatic analyzer which automatically sets a measurement reagent and sets conditions, it is possible to leave the lid open for several weeks before performing the measurement. In many cases, carbon dioxide in the atmosphere is absorbed, the pH of the AST activity measurement reagent changes, the reagent blank reaction also changes, and there is a problem that an error occurs in the obtained AST activity measurement value.

Therefore, an object of the present invention is to provide a reagent for measuring AST activity, which can suppress an increase in reagent blank reaction, that is, an increase in initial absorbance.

[0011]

[Means for Solving the Problems] The above-mentioned problems are aspartic acid containing L-aspartic acid, 2-oxoglutarate, malate dehydrogenase, lactate dehydrogenase, and reduced nicotinamide adenine dinucleotide according to the present invention. The reagent for measuring aminotransferase activity can be solved by the reagent for measuring aspartate aminotransferase activity, which further comprises a substance having an inhibitory effect on lactate dehydrogenase activity. Further, the present invention provides a test sample which may contain aspartate aminotransferase, L-aspartate, 2-oxoglutarate, malate dehydrogenase, lactate dehydrogenase, reduced nicotinamide adenine dinucleotide. And a substance having an inhibitory effect on lactate dehydrogenase activity, the method for measuring aspartate aminotransferase activity.

[0012]

BEST MODE FOR CARRYING OUT THE INVENTION The reagent for measuring AST activity of the present invention is
L-aspartic acid, 2-oxoglutaric acid, MD, L
It is an improved reagent of a known AST activity measurement reagent containing D and NADH. In a known AST activity measuring reagent containing L-aspartic acid, 2-oxoglutaric acid, MD, LD, and NADH, oxaloacetic acid produced by AST was converted to apple by MD using L-aspartic acid and 2-oxoglutaric acid as substrates. The AST activity can be measured by measuring the amount of decrease in the amount of NADH coexisted with the acid in the vicinity of the wavelength of 340 nm.

Further, in this known activity measuring reagent, as described above in the section of the prior art, in order to eliminate the influence of pyruvic acid derived from a test sample (for example, blood), for example,
In the case of a two-reagent system (for example, when at least LD is contained as the first reagent and 2-oxoglutarate is contained as the second reagent), a test sample which may contain pyruvic acid and LD The AST activity is measured without being affected by the pyruvate derived from the test sample by adding the second reagent after the pyruvate derived from the test sample is erased by mixing with a first reagent containing be able to. Even in the two-reagent system containing at least LD and 2-oxoglutaric acid as the second reagent, the pyruvic acid derived from the test sample is erased within, for example, several tens of seconds to 1 minute after the addition of the second reagent. After that, the AST activity can be measured, or even one reagent system can serve the same purpose.

The reagent for measuring AST activity of the present invention contains, in addition to these known constituents, a substance having an inhibitory action on LD activity (hereinafter referred to as LD inhibitor). The LD inhibitor used in the present invention is not particularly limited, for example, oxamic acid, oxalic acid, oxaloacetic acid, pyruvic acid, phosphoenolpyruvate, sodium dodecyl sulfate, lactic acid, or hydroxyglutaric acid, or Oxamic acid or a salt thereof (for example, a sodium salt, a potassium salt, or a lithium salt) is preferable in that these salts can be used and do not give an error to the AST activity measurement.

The concentration of the LD inhibitor contained in the reagent for measuring AST activity of the present invention varies depending on the type of LD inhibitor used, so that it does not affect the reagent for measuring AST activity.
The concentration is not particularly limited as long as it has a D activity and an LD activity that does not affect the elimination of the test sample-derived pyruvic acid, but is not particularly limited, but generally, the final concentration in the measurement system is Can be used by adjusting the content concentration in the measurement reagent so that the concentration becomes 0.001 to 100 mmol / L.

"M which does not affect the reagent for measuring AST activity
"D activity is present" means that at least the MD activity necessary for measuring the AST activity in the test sample remains at least, and even if the MD activity is inhibited by the LD inhibitor, the minimum activity is required. If the amount of MD activity remains, AST
There is no problem as an activity measuring reagent. Further, "there is LD activity that does not affect the elimination of pyruvate derived from the test sample" means that at least the LD activity that is at least necessary for eliminating the pyruvate derived from the test sample remains. This means that, even if the LD activity is inhibited by the LD inhibitor, if the minimum necessary amount of LD activity remains, it does not cause a problem as a reagent for measuring AST activity. Furthermore, since the object of the present invention is to suppress the reagent blank reaction, AST
A combination of the MD addition amount, the LD addition amount, and the LD inhibitor amount may be appropriately set so as to have the MD activity and the LD activity that do not affect the activity measurement reagent, and to minimize the reagent blank reaction. .

More specifically, for example, when oxamic acid is used as the LD inhibitor, the final concentration in the measurement system is preferably 0.005 to 5 mmol / L, more preferably 0.02 to 1 mmol / L. The expected effect can be obtained by adding the above-mentioned amount. As will be described later, when the reagent composition is divided into a first reagent and a second reagent, if the final concentration is the same as above, it is added to either one of the first reagent or the second reagent, or to both. Even if it is, the same effect can be obtained.

Among the components contained in the AST activity measuring reagent of the present invention, various components contained in known AST activity measuring reagents, namely MD, LD, NADH, L-aspartic acid and 2-oxoglutaric acid. Can be used in the same manner as a known reagent for measuring AST activity.

[0019] For example, as MD, for example, bovine heart origin, pig heart origin, Thermus flavus (Therm
us flavus), Bacillus subtilis (Bac)
illus subtilis, or Neurospora cras
A natural MD derived from sa) or a recombinant MD thereof can be used, and the origin thereof is not particularly limited. The concentration of MD contained in the reagent for measuring AST activity of the present invention is such that the final concentration in the measurement system is at least 200 U /
It can be appropriately selected and used so as to be L or more. Since the reagent for measuring AST activity of the present invention is characterized by further containing an LD inhibitor, when the MD activity is inhibited by the LD inhibitor, the inhibitory component is taken into consideration,
It can be appropriately selected and used so that the final concentration activity is at least 200 U / L or more. For example, even when oxamic acid is added as an LD inhibitor in an amount of 0.02 to 1 mmol / L, it can be appropriately selected and used so that the final concentration activity is at least 200 U / L or more. In the present specification, "MD activity"
Means the activity of reducing oxaloacetic acid to malic acid. The unit “U” is defined as the amount of enzyme activity (standard temperature is 30 ° C.) that converts 1 μmol of the substrate (oxaloacetic acid) into a product (malic acid) in 1 minute.

The LD may be, for example, derived from chicken heart, porcine heart, porcine muscle, or Leuconostost.
C. mesenteroides) -derived natural LD or a recombinant LD thereof can be used, and the origin thereof is not particularly limited. The concentration of LD contained in the AST activity measuring reagent of the present invention can be appropriately selected and used so that the final concentration in the measuring system is at least 100 U / L or more. Since the AST activity measuring reagent of the present invention is characterized by further containing an LD inhibitor, it is appropriately selected so that the final concentration activity is at least 100 U / L or more in consideration of the amount of LD inhibition by the LD inhibitor. Can be used. For example, even when oxamic acid is added as an LD inhibitor in an amount of 0.02 to 1 mmol / L, the final concentration activity is at least 100.
It can be appropriately selected and used so as to be U / L or more. In the present specification, the “LD activity” means the activity of reducing pyruvic acid to lactic acid. The unit “U” is 1 μmol of substrate (pyruvic acid) per minute.
Is defined as the amount of enzyme activity (standard temperature is 30 ° C.) that converts the product into a product (lactic acid).

Regarding the concentration of NADH contained in the AST activity measuring reagent of the present invention, the final concentration in the measuring system is preferably 0.05 to 2 mmol / L, more preferably 0.1 to 2.
It can be used in a range of 0.5 mmol / L. Regarding the concentration of L-aspartic acid as a substrate, the final concentration in the measurement system is preferably 100 to 3000 mmol.
/ L, and more preferably in a range of 200 to 2000 mmol / L. The concentration of 2-oxoglutaric acid, which is another substrate, can be used within the range where the final concentration in the measurement system is preferably 1 to 500 mmol / L, more preferably 5 to 100 mmol / L.

The reagent for measuring AST activity of the present invention is a known A
Similar to the ST activity measurement reagent, a suitable buffer may further be contained. As the buffer, a conventionally known buffer may be appropriately selected and used as long as it does not adversely affect the AST activity measurement. . Specifically, for example, tris (hydroxymethyl) aminomethane, phosphoric acid, 2- [4- (2
-Hydroxyethyl) -1-piperazyl] ethanesulfonic acid, bis (2-hydroxyethyl) iminotris (hydroxymethyl) methane, 2-hydroxy-N-tris (hydroxymethyl) methyl-3-aminopropanesulfonic acid, N-tris (Hydroxymethyl) methyl-2-
Aminoethanesulfonic acid, n-ethylmorpholine or the like can be used. Furthermore, the AST activity measuring reagent of the present invention comprises, in addition to the above-mentioned essential components and buffers, components that are generally added, if necessary, such as chelating agents [eg, ethylenediaminetetraacetic acid (EDTA), etc.],
An antiseptic (eg, azide), a stabilizer (eg, albumin or glycerol), and / or various surfactants can be added as appropriate.

The reagent composition of the AST activity measuring reagent of the present invention is not particularly limited in the same manner as the known AST activity measuring reagent containing LD, and it may be combined into one reagent, or It is also possible to have a reagent configuration of two or more reagents. For example, from the viewpoint of eliminating the influence of pyruvic acid derived from the test sample, a reagent containing at least LD (for example, a first reagent in the case of a two-reagent system) and a test sample after the reagent As a reagent to be brought into contact with (for example, a second reagent in the case of a two-reagent system), a reagent composition of two or more reagents (especially two-reagent system) containing a reagent containing at least 2-oxoglutarate. However, the reagent composition of the AST activity measuring reagent of the present invention is not limited to this. Further, in general, it is preferable to divide each of the constituent components into stable conditions so that the reagents can be reacted under the optimal conditions for activity measurement. Such reagent composition is divided into, for example, a first reagent containing alkaline and stable NADH, MD, LD, etc. and a second reagent containing 2-oxoglutarate, etc., and is optimal for activity measurement during the reaction. It is possible to configure each of these reagents to have a reagent concentration and a pH, and further add an LD inhibitor and L-aspartic acid to either or both of the first reagent and the second reagent. ,
It is not limited to this.

The reagent for measuring AST activity of the present invention is, for example,
It can be used for the AST activity measuring method of the present invention. In the method for measuring AST activity of the present invention, a test sample that may contain AST is contacted with L-aspartic acid, 2-oxoglutarate, MD, NADH, and an LD inhibitor to reduce the amount of NADH. Is measured at a wavelength of around 340 nm, whereby the AST activity can be measured. The test sample is not particularly limited as long as it is a test sample that may contain AST activity,
For example, a biological fluid commonly used for clinical diagnosis, such as blood, serum, plasma, or urine, or cell tissue,
Or an experimental sample etc. can be mentioned.

[0025]

The present inventors believe that the reason why the reagent blank reaction is suppressed in the AST activity measuring reagent of the present invention is due to the following action. The present invention is not limited to the following reasoning. LD contained in the AST activity measuring reagent of the present invention has an activity of reducing 2-oxoglutarate, that is, 2-hydroxyglutarate dehydrogenase (HGD) activity, in addition to dehydrogenase activity for lactic acid which is a main enzyme activity. [Clinical Chemistry, 18 (4), 250-262 (1989)].
If this reaction is shown, it is as follows.

As described above in the section of the prior art, the commonly used AST activity measurement including the recommended method published by the Japan Society for Clinical Chemistry uses 2-oxoglutaric acid as a substrate, and therefore LD is 2-oxoglutaric acid. Having a dehydrogenase activity (that is, HGD activity) for acid, NADH
To oxidize and increase the reagent blank and reagent blank reaction. In addition, the dehydrogenase activity for 2-oxoglutarate changes due to the change in pH, resulting in an error in the measured AST activity. For example, in an analyzer called an automatic analyzer that automatically sets a measurement reagent and sets the conditions, the analyzer is often left standing with the lid open for several weeks. The carbon dioxide in the sample is absorbed, the pH of the AST activity measurement reagent changes, the reagent blank reaction also changes, and an error occurs in the obtained AST activity measurement value. The reagent blank reaction occurring here is considered to be a reaction for reducing 2-oxoglutarate shown by LD itself, that is, HGD activity.

The present inventor confirmed that HG
D activity is present, and the reagent blank also changes depending on the amount of LD added to the AST activity measurement reagent. Also, HG
The D activity has an optimum pH from weakly acidic to around neutral. For example, if the LD amount is increased, the reagent blank also becomes large, and when measuring AST activity under weak alkaline conditions, the reagent blank becomes large due to the decrease in the reagent pH after opening and storing the reagent container, and accurate AST activity measurement You can't get the value. To reduce the reagent blank, it is conceivable to reduce the LD amount. However, if the LD amount is too small, the LD amount necessary for eliminating the pyruvic acid derived from the test sample will be insufficient and quantitative determination will be obtained. I will not be able to. In addition, the reagent blank rise due to the decrease in the reagent pH after opening and storing the reagent container can avoid the reagent blank by shifting the pH during AST activity measurement to alkaline, but the optimum pH for AST activity is weakly alkaline. , It's not an appropriate method.

Conventionally, no inhibitor of HGD activity has been known at all, and under such circumstances, the present inventor has made known AST.
By adding an LD inhibitor to the activity measurement reagent, LD
It is considered that the HGD activity shown by itself was inhibited, and as a result, it was possible to reduce the reagent blank without impairing the quantification property and to avoid the reagent blank increase due to the decrease in the reagent pH after the reagent container was opened and stored. There is.

[0029]

The present invention will be described in detail below with reference to examples, but these do not limit the scope of the present invention.

Preparation Example 1 As a reagent for measuring AST activity of the present invention,
A two-reagent system reagent consisting of a first reagent having the composition shown in Table 1 and a second reagent having the composition shown in Table 2 was prepared. As a conventionally known AST activity measuring reagent for comparison, a two-reagent system reagent having the same composition as the first reagent and the second reagent except that the first reagent did not contain oxamic acid was prepared. Hereinafter, the two-reagent type reagent for comparison is referred to as "reagent A", and the two-reagent type reagent of the present invention is referred to as "reagent B". The prepared Reagent A and Reagent B were each placed in a sealable container and stored in a sealed state until they were used in the following evaluation examples.

<< Table 1 >> Concentration Ingredients 20 mmol / L Tris-hydrochloric acid (pH 9.20) 135 mmol / L L-Aspartic acid 0.5 mmol / L Oxamic acid 0.095% Sodium azide 0.25 mmol / L NADH 3KU / L LD (recombinant LD; manufactured by Oriental Yeast Co., Ltd.) 2KU / L MD (manufactured by Amano Enzyme Inc.)

<< Table 2 >> Concentration Ingredients 272 mmol / L Tris-hydrochloric acid (pH 4.50) 425 mmol / L L-aspartic acid 42 mmol / L 2-oxoglutarate 0.01% EDTA2Na

[0032]

[Evaluation Example 1] << Evaluation of AST activity measuring reagent of the present invention >> (1) Measurement of reagent blank First reagent 60 mL and second reagent 20 mL of the comparative reagent A prepared in Preparation Example 1 above, and the present invention Reagent B
60 mL of the first reagent and 20 mL of the second reagent of No. 1 were filled in reagent containers for an automatic analyzer (7170S; manufactured by Hitachi, Ltd.), and these reagent containers were set in the automatic analyzer and left for 5 weeks. . The automatic analyzer under evaluation has a cooling device attached to the portion where the reagent container is set and is kept at about 10 ° C. The lid of the reagent container was left open. Immediately after standing, after 1 week, 2 weeks, 3 weeks, 4 weeks, and 5 weeks, the reagent blanks of reagent A and reagent B were measured according to the following procedures.

More specifically, after adding 7.5 μL of physiological saline as a sample to the reaction cell of the automatic analyzer, 150 μL of the first reagent was added and stirred, and after heating at 37 ° C. for 5 minutes , 50 μL of the second reagent was added and stirred, and the mixture was further heated at 37 ° C. for 5 minutes. About 1 after adding the second reagent
The amount of change in absorbance at a wavelength of 340 nm was measured from the lapse of minutes until the lapse of 5 minutes (that is, 4 minutes), and the amount of change in absorbance per minute was calculated as the blank sensitivity. The results are shown in Fig. 1. As shown in FIG. 1, when the lid is left open, the reagent blank of the comparative reagent A rises with time, whereas the reagent blank of the reagent B of the present invention has a reagent blank. Changed little.

(2) Measurement of AST activity in pooled serum In the same manner as in Evaluation Example 1 (1) above, Reagent A and Reagent B prepared in Preparation Example 1 were left for 5 weeks. Immediately after leaving, 1
After a lapse of weeks, 2 weeks, 3 weeks, 4 weeks, and 5 weeks, the AST activity in the sample (pool serum) was measured according to the following procedures. That is, pooled serum or physiological saline (reagent blank)
Add 150 μL of the first reagent to 7.5 μL and stir,
After heating at 0 ° C for 5 minutes, 50 µL of the second reagent was added and stirred, and further heated at 37 ° C for 5 minutes. The amount of change in absorbance at a wavelength of 340 nm from about 1 minute to 5 minutes after the addition of the second reagent (that is, 4 minutes) is measured,
The amount of change in absorbance per minute was calculated. The ATL activity was converted based on the measurement results obtained using an enzyme calibrator (manufactured by Kokusai Reagent Co., Ltd.) instead of pooled serum.

The results are shown in FIG. As shown in FIG. 2, with respect to the measured value of pooled serum, when it is left with the lid open, the measured value of AST activity of the reagent A for comparison increases with time. In the reagent B of the present invention,
Almost no variation was observed in the measured AST activity.

[0036]

[Evaluation Example 2] << L in the reagent for measuring AST activity of the present invention
Evaluation on Stability of D >> In this evaluation example, 60 mL of the first reagent of the reagent A for comparison prepared in Preparation Example 1 and 60 mL of the first reagent of the reagent B of the present invention were each prepared to 70%.
It is sealed in a polyethylene bottle with a lid of mL volume and stored at 25 ° C or 37 ° C for 3 weeks. LD contained in each first reagent
The change with time in the residual rate of activity was examined. Specifically, immediately after storage, and after 1 week and 2 weeks from the start of storage,
And after 3 weeks, each reagent was collected and each reagent contained 0.1% bovine serum albumin in 50 mm at the time of measurement.
A sample diluted with 1/10 concentration of ol / L phosphate buffer (pH 7.50) was used as a sample, and the analysis was performed using an automatic analyzer (7170S; manufactured by Hitachi, Ltd.) according to the following procedure.

7.5 μL of the sample was added to the LD activity measuring reagent 1
[50 mmol / L phosphate buffer (pH 7.50) and 0.25 mmol / L-NADH] 150 μL was added,
After heating at 37 ° C for 5 minutes, LD activity measurement reagent 2 [50
mmol / L phosphate buffer (pH 7.50) and 12 m
mol / L lithium pyruvic acid salt] 30 μL was added and stirred, and further heated at 37 ° C. for 5 minutes. The amount of change in absorbance per minute at a wavelength of 340 nm from about 1 minute to 2 minutes after the addition of the LD activity measuring reagent 2 was measured. LD with the change in absorbance on the first day of storage as 100%
The residual rate of activity was calculated.

The results are shown in FIG. In FIG. 3, the symbols “1W”, “2W”, and “3W” mean the results after 1 week, 2 weeks, and 3 weeks from the start of storage, respectively. As shown in FIG.
Under both conditions of 7 ° C. and 25 ° C., the residual rate of LD activity was higher in the reagent B of the present invention than in the comparative reagent A.

[0039]

According to the AST activity measuring reagent of the present invention,
Since an increase in the reagent blank reaction, that is, an increase in initial absorbance can be suppressed, an accurate AST activity measurement value can be obtained. In addition, it has a stabilizing effect on the LD.

[Brief description of drawings]

FIG. 1 is a graph showing changes in blank sensitivity with time in the present invention and a comparative AST activity measurement reagent.

FIG. 2 is a graph showing changes over time in AST activity measurement values in pool serum, which were measured using the AST activity measurement reagent of the present invention and the comparative AST activity measurement reagent.

FIG. 3 is a graph showing LD stability in the present invention and a comparative AST activity measurement reagent.

Claims (3)

[Claims] 1. A lactate dehydrogenase activity in a reagent for assaying aspartate aminotransferase activity containing L-aspartate, 2-oxoglutarate, malate dehydrogenase, lactate dehydrogenase, and reduced nicotinamide adenine dinucleotide. A substance having an inhibitory effect on
The aspartate aminotransferase activity measuring reagent, which further comprises: 2. The reagent for measuring aspartate aminotransferase activity according to claim 1, wherein the substance having an inhibitory effect on lactate dehydrogenase activity is oxamic acid or a salt thereof. 3. A test sample which may contain aspartate aminotransferase, L-aspartate, 2-oxoglutarate, malate dehydrogenase, lactate dehydrogenase, reduced nicotinamide adenine dinucleotide, And a substance having an inhibitory action on lactate dehydrogenase activity are brought into contact with each other, and a method for measuring aspartate aminotransferase activity.