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Reagent for determining adenosine deaminase and preparing method thereof

An adenosine deaminase and reagent technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., and can solve problems such as difficult enzyme content determination.

Inactive Publication Date: 2003-11-19
TECOM SCI CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It is difficult to determine the enzyme content in body fluid samples by this method in practice

Method used

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Examples

Experimental program
Comparison scheme
Effect test

specific Embodiment Embodiment 1

[0017] Specific Examples Example 1: Prepare the reagent for measuring adenosine deaminase, which is composed of the following components according to the content per liter,

[0018] Reagent 1:

[0019] Tris buffer 100mM, (50-200mM), pH8.2, 37°C,

[0020] a-ketoglutarate 7.5mM, (2-20mM)

[0021] Bovine serum albumin 5g / L (1-10g / L),

[0022] Adenosine diphosphate potassium salt 2mM (0.2-20mM)

[0023] Glutamate dehydrogenase (from bovine liver) 3500U / L (500-10000U / L)

[0024] NAD0.6mM (0.15-1.2mM)

[0025] D-glucose 40mM (10-200mM)

[0026] Glucose-6-phosphate dehydrogenase (Leuconostoc) 3700U / L (500-10000U / L)

[0027] Potassium dihydrogen phosphate 5mM (2-100mM)

[0028] EDTA.Na 2 .2H 2 O1.5Mm (0.2-20mM)

[0029] Sodium azide 7.0mM (1-10mM)

[0030] According to the method for preparing the reagent solution above, after the preparation of the above reagent is completed, place it at 2-8°C for 1-7 days, and the reagent blank absorbance is ≥1.20, 340m, 10mm optical path...

Embodiment 2

[0040] According to the method for preparing the reagent solution above, configure reagent 2 and store it at 2-8°C. When measuring samples, there is a two-point method, the ratio of reagent 1: sample: reagent 2 is 200:20:100, the temperature is 37°C, reagent 1 is added to the sample or calibrator and incubated at the measurement temperature for 300 seconds to remove the ammonia in the sample , add reagent 2 to start the measurement, the delay time is 0 seconds, the measurement time is 300 seconds, and the reading is selected from 2 valid points within the measurement time. Embodiment 2: total adenosine deaminase liquid single reagent

[0041] Tris buffer 100mM, (50-200mM), pH7.4, 37°C,

[0042] a-ketoglutarate 7.5mM, (1-20mM)

[0043] Bovine serum albumin 5g / L (1-10g / L),

[0044] Adenosine diphosphate potassium salt 2.5mM (0.5-20mM)

[0045] Adenosine 2mM (0.2-20mM)

[0046] Glutamate dehydrogenase (beef liver source) 4500U / L (500-10000U / L)

[0047] NADP0.45mM (0.15-1.2m...

Embodiment 3

[0053] According to the method for preparing the reagent solution above, configure reagent 2 and store at 2-8°C for 1-7 days. Reagent blank absorbance ≥ 1.20, 340nm, 10mm optical path. When measuring samples, adopt the two-point method, reagent 1:15, temperature 37°C, delay time 0 seconds, remove ammonia in the sample, measure time 300 seconds, select 2 valid points for reading within the measure time. Example 3: Adenosine deaminase isozyme isozyme 2 liquid double reagent

[0054] Reagent 1:

[0055] Tris buffer 100mM, (50-200mM), pH8.2, 37°C,

[0056] a-ketoglutarate 7.5mM, (2-20mM)

[0057] Bovine serum albumin 5g / L (1-10g / L),

[0058] Adenosine diphosphate potassium salt 2mM (0.2-20mM)

[0059] Glutamate dehydrogenase (from bovine liver) 3500U / L (500-10000U / L)

[0060] NADP0.45mM (0.15-1.2mM)

[0061] EHNA0.15mM (0.05-0.5mM)

[0062] D-glucose 40mM (10-200mM)

[0063] Glucose-6-phosphate dehydrogenase (Leuconostoc) 3700U / L (500-10000U / L)

[0064] Potassium dihydro...

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PUM

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Abstract

A reagent for detecting the activity of adenosine deaminase and its isozymes in body fluid specimen by enzyme method is prepared from the oxidation-type micotinamide coenzyme or its analog, and substrate to generate the slow-reaction enzyme-substrate-DNA (or NADP) system.

Description

technical field [0001] The invention relates to a reagent for measuring adenosine deaminase and a preparation method thereof. Background technique [0002] Adenosine deaminase is an important enzyme in the metabolism of purine nucleosides and a nucleic acid metabolizing enzyme that has an important relationship with the immune activity of the body's cells. It is widely distributed in various tissues of the human body, with the highest content in thymus, spleen and other lymphoid tissues. There are three isozymes of adenosine deaminase, namely adenosine deaminase 1, adenosine deaminase 1+cp and adenosine deaminase 2. In recent years, the determination of the activity levels of adenosine deaminase and its isoenzymes in blood, body fluids and other samples for the diagnosis, differential diagnosis, treatment and immune function of certain diseases has received more and more clinical attention. The lack of adenosine deaminase activity is related to severe combined immunodefici...

Claims

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Application Information

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IPC IPC(8): C12Q1/25C12Q1/32
Inventor 颜箫
Owner TECOM SCI CORP
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