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N-acetyl-beta-D glucosidase reagent and detection method

A glucosidase and detection method technology, applied in the field of N-acetyl-β-D glucosidase detection reagents, can solve the problems of long operation time, cumbersome operation, insoluble substrate, etc., and achieve improved analysis sensitivity and stability Strong, good water solubility

Inactive Publication Date: 2016-03-23
BIOBASE BIODUSTRY (SHANDONG) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This substrate is cheap and easy to obtain, but the sample blank must be set, and the operation time is long, so it is not suitable for large-scale automated analysis
The CNP-NAG method uses 2-chloro-4-nitrophenylacetylglucosaminidide as the substrate, and the product after NAG enzymolysis can be directly colored without alkalization, and it does not need to set a sample blank, which can realize large-scale automation operation, but the substrate is insoluble, the pH value of the buffer needs to be strictly controlled during the determination, and the substrate solution should be freshly prepared, and the operation is cumbersome

Method used

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  • N-acetyl-beta-D glucosidase reagent and detection method
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  • N-acetyl-beta-D glucosidase reagent and detection method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Detection reagents for N-acetyl-β-D-glucosidase, including reagent R1 and reagent R2:

[0034] 1) The components of reagent R1 are:

[0035] Citric acid-sodium hydrogen phosphate disodium buffer solution (PH=5.0)..........................100mmol / L

[0036] VRA-NAG................................................ ...................................3mmol / L

[0037] BSA................................................................ ...................................1g / L

[0038] Ethylene glycol................................................ ...................................1ml / L

[0039] TritonX-100................................................................ ...................................1ml / L

[0040] Ascorbate oxidase................................................ ...................1ml / L

[0041] Preservative NaN 3. ................................................... ..............2mmol / L

[0042] 2) The components of reagent R2 are:

[0043] ...

Embodiment 2

[0051] Interference test: Take fresh urine, divide it into 2 equal parts, then divide each equal part into 5 equal parts, add different interfering substances, and make the concentration in the urine meet the requirements in Table 2. Then, the reagents obtained in Example 1 were used to compare and measure the UA content in serum at the same time as the common and recognized NAG reagents in the market. Relative deviation (%) = (measuring mean value of interference samples - measuring mean value of control samples) / measured mean value of control samples × 100%.

[0052] As can be seen from Table 2, the reagents of Example 1 are lactic acid≤0.9g / L, pyruvic acid≤20mg / L, tartaric acid≤17mg / L, uric acid≤g / L, glucose≤g / L, bilirubin≤mg / L L did not significantly interfere with the determination results. However, the contrast reagent was significantly interfered when the interfering substance at the above-mentioned concentration existed, which shows that the anti-interference performa...

Embodiment 3

[0056] Correlation test: using the formula in Example 1 to prepare reagents, and compared with the NAG kit of a company approved by the State Food and Drug Administration, which is common in the market, and tested 20 clinical samples at the same time, the test results are shown in Table 3. And obtained the correlation curve of the two reagents (such as figure 1 Shown), the results show that the correlation coefficient of the two kits is 0.9994, indicating that there is a great correlation between the two.

[0057] Table 3 Example 1 and the market common and recognized NAG assay kit comparative detection results

[0058]

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Abstract

The invention relates to the field of reagent NAG detection technologies, in particular to an N-acetyl-beta-D glucosidase reagent. The reagent R1 contains a buffer solution, VRA-NAG, bovine serum albumin, TritonX-100, ethylene glycol, ascorbic acid oxidase and a preservative. The reagent R2 contains a buffer solution and a preservative. The reagent adopts a novel substrate VRA-NAG, the substrate has better solubility in water compared with other substrates, is good in stability and is a reliable NAG substrate. The reagent uses citric acid-disodium hydrogen phosphate as the buffer solution, the buffering capacity of the reagent is greatly improved, and meanwhile a reaction system is not destroyed. The added TritonX-100 and the ethylene glycol serve as cosolvents to promote dissolution of the substrate and have an obvious solubilization assisting effect. BSA is added to serve as a stabilizer, and the stability and anti-interference capability of the reagent are greatly enhanced. The N-acetyl-beta-D glucosidase reagent is simple in configuration and low in price and is very suitable for large-area clinic popularization.

Description

technical field [0001] The invention relates to the technical field of N-acetyl-β-D glucosidase detection, in particular to a detection reagent for N-acetyl-β-D glucosidase, and also relates to a detection method using the detection reagent. Background technique [0002] N-acetyl-β-D-glucosaminidase (NAG) is a hydrolytic enzyme present in the lysosome of cells, which is widely distributed in various tissues of the human body, but the highest content is in the prostate and proximal renal tubules of the kidney. The molecular weight of NAG is about 130-140KD. Under normal circumstances, NAG in serum cannot be excreted from urine through glomeruli. NAG in urine mainly comes from the epithelial cells of the renal proximal tubule, but the content in the urine of healthy people is very small. The increase of urinary NAG activity level is a sensitive indicator of early kidney injury and diabetic microangiopathy. NAG activity can be used for early diagnosis of tubulointerstitial ne...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/573
CPCG01N33/573
Inventor 谭柏清刘慧王绮赵新
Owner BIOBASE BIODUSTRY (SHANDONG) CO LTD
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