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Method for detecting 1,5-anhydro sorbitol and related diagnostic kit

A technology of sorbitan and reagent kits, which is applied in the field of clinical biochemical diagnosis, can solve the problems of high cost of reagent kits, and achieve the effects of ensuring stability, convenient use, and strong removal ability

Active Publication Date: 2011-08-17
BEIJING STRONG BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] However, the cost of the kits used in the prior art is high, and further improvement is still needed in terms of stability, sensitivity, and specificity. Therefore, it is necessary to improve the detection method of 1,5-sorbitol and related diagnostic kits to Meets requirements for clinical testing and diagnostics, as well as chemical analysis

Method used

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  • Method for detecting 1,5-anhydro sorbitol and related diagnostic kit
  • Method for detecting 1,5-anhydro sorbitol and related diagnostic kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] Click on the ingredients and dosage to configure the 1,5-AG diagnostic kit. In the present invention, unless otherwise specified, percentages are mass-volume ratios.

[0091] Reagent 1

[0092] Phosphate buffer 100mM

[0093] Adenosine diphosphate 5mM

[0094] Magnesium Chloride 5Mm

[0095] ADP-dependent hexokinase 20U / ml

[0096] Tio-NAD 1g / L

[0097] Sodium azide 0.5g / L

[0098] Reagent 2

[0099] Tris buffer 10mM

[0100] 1,5-anhydroglucitol-6-phosphate dehydrogenase 10U / ml

[0101] NADH 6g / L

[0102] Sodium azide 0.5g / L

[0103] Set on the automatic biochemical analyzer (Olympus-400): reaction temperature 37°C, reaction time 1.5 minutes, test wavelength 405nm, test sub-wavelength 540nm. The ratio of the tested potassium ion sample to the reagent is: sample: reagent 1: reagent 2 = 5 μl: 180 μl: 60 μl, and the reaction is a drop reaction.

[0104] First add the sample and reagent 1, the two are automatically mixed in the automatic biochemical analyzer, dete...

Embodiment 2

[0107] According to Embodiment 1, a sugar removal system is added to configure a 1,5-AG detection kit.

[0108] Reagent 1

[0109] Tris-HCl 100mmol / L

[0110] Adenosine diphosphate 5mmol / L

[0111] Adenosine triphosphate 5mmol / L

[0112] Hexokinase 5U / ml

[0113] Glucose-6-phosphate 5U / ml

[0114] Hexose phosphate isomerase 10U / ml

[0115] Magnesium chloride 5mmol / L

[0116] ADP-dependent hexokinase 20U / ml

[0117] Tio-NAD 1g / L

[0118] Sodium azide 0.5g / L

[0119] Reagent 2

[0120] Tris buffer 10mM

[0121] 1,5-anhydroglucitol-6-phosphate dehydrogenase 10U / ml

[0122] NADH 6g / L

[0123] Sodium azide 0.5g / L

[0124] Using the experimental method of Example 1, the sugar removal system was tested.

[0125] sample

[0126] 1,5-AG pure product solution (150umol / l)

[0127] According to the above data, the sugar removal system can remove the interference of glucose on the detection, but there is still some interference of glucose on the system.

Embodiment 3

[0129] According to Embodiment 1, a sugar removal system is added to configure a 1,5-AG detection kit.

[0130] Reagent 1

[0131] Tris-HCl 100mmol / L

[0132] Adenosine diphosphate 5mmol / L

[0133] Adenosine triphosphate 5mmol / L

[0134] Glucokinase 5U / ml

[0135] Glucose-6-phosphate 5U / ml

[0136] Hexose phosphate isomerase 10U / ml

[0137] Magnesium chloride 5mmol / L

[0138] ADP-dependent hexokinase 20U / ml

[0139] Tio-NAD 1g / L

[0140] Sodium azide 0.5g / L

[0141] Reagent 2

[0142] Tris buffer 10mM

[0143] 1,5-anhydroglucitol-6-phosphate dehydrogenase 10U / ml

[0144] NADH 6g / L

[0145] Sodium azide 0.5g / L

[0146] Using the experimental method of Example 1, the sugar removal system was tested.

[0147] sample

[0148] According to the above data, the sugar removal system can remove the interference of glucose on the detection, which is obviously stronger than that of Example 3.

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Abstract

The invention relates to a method for detecting 1,5-anhydro sorbitol and a related diagnostic kit. In the method, a sample and adenosine diphosphate are reacted in the presence of adenosine diphosphate dependent hexokinase serving as a catalyst to form anhydroglucose-6-phosphoric acid and adenylic acid, the anhydroglucose-6-phosphoric acid and thio-oxidative NAD(nicotinamide adenine dinucleotide) are reacted in the presence of 1,5-AG(anhydro glucitol)-6-phosphate dehydrogenase(PDH) serving as a catalyst to form thio-reduced coenzymes and compounds of C6H11O8P, and simultaneously the generated C6H11O8P is reacted in the presence of AG-6-PDH and reduced NAD to form AG-6-P and oxidative coenzymes; and because of the cyclic amplification, the sensitivity of 6-anhydro sorbitol is improved. The invention also provides a detection kit for 1,5-anhydro sorbitol based on the method. The kit is convenient and easy to use, can quickly finish detection on a common ultraviolet / visible light analyzer or semi- / full-automatic biochemical analyzer, does not need special or additional instruments and is low in cost.

Description

technical field [0001] The invention relates to the field of clinical biochemical diagnosis, and relates to a detection method for enzymatic detection of 1,5-sorbitol and a related diagnostic kit. Further, the present invention relates to a method and a kit for detecting 1,5-sorbitol in blood by circulating enzyme method. Background technique [0002] 1,5-anhydroglucitol (1,5-AG), also known as 2-deoxyglucose, 1,5-anhydroglucitol, is a six-carbon monosaccharide with a pyran ring structure. It is the reduced form of glucopyranose (the lack of a hydroxyl group at the C-1 position), forming a C1 chair-shaped oxygen ring at the pyran position, and the molecular formula is C 6 h 12 o 5 (Molecular weight 164.2, CAS number 154-58-5), the structure is very similar to that of glucose, the main difference between the two is that the hydroxyl group at the C-1 position of glucose is replaced by hydrogen, and this substitution is exactly 1,5- Sorbitan is the chemically stable base. ...

Claims

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Application Information

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IPC IPC(8): C12Q1/48C12Q1/32
Inventor 刘瑶杜爱铭刘希蔡华雅
Owner BEIJING STRONG BIOTECH
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