CN115710597A - Urea detection kit - Google Patents
Urea detection kit Download PDFInfo
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- CN115710597A CN115710597A CN202211336926.9A CN202211336926A CN115710597A CN 115710597 A CN115710597 A CN 115710597A CN 202211336926 A CN202211336926 A CN 202211336926A CN 115710597 A CN115710597 A CN 115710597A
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Abstract
The invention provides a urea detection kit, which belongs to the technical field of medical inspection and comprises a reagent 1, a reagent 2, a calibrator and a quality control product. The main components in the reagent 1 are as follows: buffer solution, alpha-ketoglutaric acid, glutamate dehydrogenase, reduced coenzyme and stabilizer. The main components in the reagent 2 are as follows: buffer solution, alpha-ketoglutaric acid, urease and bovine serum albumin. According to the invention, ADP and GDP are added into the reagent 1 to increase the activity of glutamate dehydrogenase, so that the detection performance of the reagent is improved.
Description
Technical Field
The invention belongs to the field of biological detection, and particularly relates to a urea detection kit.
Background
Urea is one of the end products of amino acid catabolism in the human body. The urea in blood comes from liver and is excreted with urine through kidney. Various kidney diseases and liver diseases can be diagnosed by detecting the content of urea in blood.
At present, the detection of urea in blood mainly adopts a urease-glutamic acid dehydrogenase method, and the diagnostic reagent can be directly applied to a biochemical analyzer to detect the urea content in serum, so that the method has the advantages of rapidness and accuracy.
The reaction principle is as follows: urea generates ammonium ions under the action of urease, the ammonium ions, alpha-ketoglutaric acid and reduced coenzyme I generate glutamic acid and oxidized coenzyme I under the catalysis of glutamate dehydrogenase, and the reduction rate of the absorbance of the reduced coenzyme I is measured at 340nm and is in direct proportion to the content of urea in a sample. The activity of glutamate dehydrogenase directly influences the reaction result, and the reduction of the activity of glutamate dehydrogenase in the using process can cause the performance reduction of the reagent and influence the detection result.
Disclosure of Invention
Aiming at the technical problems, the invention aims to provide a urea detection kit with stable detection performance.
In order to achieve the above object, the present invention provides the following technical solutions:
a urea detection kit comprises a reagent 1, a reagent 2, a calibrator and a quality control product, wherein the reagent 1 mainly comprises the following components: tris-succinate buffer (pH9.6), alpha-ketoglutarate, glutamate dehydrogenase (GLDH) allosteric activator, reduced coenzyme I (NADH), and the main components of the reagent 2 comprise Tris buffer (pH7.8), alpha-ketoglutarate, urease and Bovine Serum Albumin (BSA).
Further, in the reagent 1: tris-succinate buffer (pH9.6) concentration 100-200mmol/L, alpha-ketoglutarate concentration 5.0-10.0mmol/L, glutamate dehydrogenase (GLDH) concentration 0.5-3.0KU/L, reduced coenzyme I (NADH) concentration 0.1-0.5mmol/L.
Further, in the reagent 1, the optimal selection concentration of Tris-succinate buffer solution (pH9.6) is 115mmol/L, the optimal selection concentration of alpha-ketoglutarate is 8mmol/L, the optimal selection concentration of glutamate dehydrogenase (GLDH) is 0.8KU/L, and the optimal selection concentration of reduced coenzyme I (NADH) is 0.35mmol/L.
Further, in the reagent 2: tris buffer (pH 7.8) concentration: 40-150mmol/L, concentration of alpha-ketoglutaric acid: 5-15mmol/L, urease (URH) concentration: 20-50KU/L, bovine Serum Albumin (BSA) concentration: 0.8-1.5g/L.
Further, in the reagent 2: the optimal selection concentration of Tris buffer (pH7.8) is 115mmol/L, the optimal selection concentration of alpha-ketoglutaric acid is 8mmol/L, the optimal selection concentration of Urease (URH) is 40KU/L, and the optimal selection concentration of Bovine Serum Albumin (BSA) is 1g/L.
Further, the glutamate dehydrogenase (GLDH) allosteric activator is one or two of Adenosine Diphosphate (ADP) and Guanine Dinucleotide Phosphate (GDP).
Further, the concentration of the glutamate dehydrogenase (GLDH) allosteric activator is 0.1mg/L-5mg/L.
The kit is used for detecting the urea amount in human serum, and is stored in a dark place at the temperature of 2-8 ℃, and the effective period is 18 months; after the reagent is unpacked, the reagent is stored in a dark place at the temperature of 2-8 ℃, and the accuracy and precision of the reagent are better than those of the reagent without the stabilizer within 60 days.
The invention has the beneficial effects that:
1. the glutamic acid dehydrogenase activator is added into the reagent 1, so that the performance index of the reagent is effectively improved. The detection accuracy and precision performance of the reagent within 60 days after the cover is opened are superior to those of a control group.
Detailed Description
The invention is further illustrated by the following examples, but is not to be construed as being limited thereby.
Example 1
The invention provides a urea detection kit, which comprises the following main components:
the test conditions are as follows: the reaction temperature is 37 ℃, the detection wavelength is 340nm, and the ratio of a reagent sample to a reagent sample R1: r2: s = 150.
Accuracy: the relative deviation calculation was performed by measuring the kit with a certified reference substance having a concentration value of 4.57mmol/L, and repeating the measurement 3 times.
M-mean test result; t-certified reference substance index value.
Precision: and (4) repeatedly testing the high-value and low-value quality control products for 10 times, and calculating the SD and CV values.
According to the requirement of a row standard YY/T1201-2013 urea nitrogen detection kit, the relative accuracy deviation is not more than +/-15%; precision is not greater than 5.0%.
The accuracy and precision of the reagents after calibration with the calibrator were measured at 0, 10, 20, 30, 40, 50, and 60 days, respectively, and the results are as follows.
Accuracy of
Precision degree
The test result shows that: the accuracy of the reagent in the embodiment 1 is better than that of the control group within 60 days after uncovering, the deviation of the accuracy of the embodiment 1 in 60 days is 1.09%, the deviation of the control group is 5.18%, the data are in the range of the standard requirement, but the deviation of the control group is larger than that of the embodiment 1. The precision values of example 1 were 1.53% and 1.34% over 60 days, respectively. The precision values of the control groups were 5.60% and 3.76%, respectively. The comparison group is larger than the embodiment 1, and the measured low-value quality control CV value of the comparison group exceeds the specified value of the row standard.
Example 2
The invention provides a urea detection kit, which comprises the following main components:
the test conditions were the same as in example 1, and the accuracy and precision of example 2 and the control group without ADP addition were determined on days 0, 10, 20, 30, 40, 50 and 60 after the reagents were calibrated with the calibrator. And (3) test results: accuracy and precision increase as decap time increases. Accuracy: the data are within the range required by the standard, the deviation of the accuracy of the example 2 is 1.03 percent at 60 days, the deviation of the control group is 5.37 percent, and the accuracy and precision of the example 2 are better than those of the control group. Precision: the precision values of example 1 were 1.47% and 1.45% over 60 days, respectively. The measured values of the control group precision are 5.76 percent and 3.94 percent respectively, the control group is larger than the example 2, and the measured value of the control group low quality control CV value exceeds the standard value.
Example 3
The invention provides a urea detection kit, which comprises the following main components:
the test conditions were the same as in example 1, and the accuracy and precision of example 3 and the control group without GDP added were measured at 0, 10, 20, 30, 40, 50, and 60 days, respectively, after the reagents were calibrated with the calibrator. And (3) test results: accuracy and precision increase as decap time increases. Accuracy: the data are in the range of the row standard, the deviation of the accuracy of the example 2 is 1.23 percent at 60 days, the deviation of the control group is 5.17 percent, and the accuracy and the precision of the example 3 are better than those of the control group. Precision: the precision values of example 1 were 1.52% and 1.37%, respectively, measured over 60 days. The control group precision was 5.61% and 3.69%. The control group is larger than example 3, and the control group determines that the low-value quality control CV value exceeds the standard value.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.
Claims (7)
1. A urea detection kit is characterized by comprising a reagent 1, a reagent 2, a calibrator and a quality control product, wherein the reagent 1 mainly comprises the following components: tris-succinate buffer (pH9.6), alpha-ketoglutarate, glutamate dehydrogenase (GLDH) allosteric activator, reduced coenzyme I (NADH), and the main components of the reagent 2 comprise Tris buffer (pH7.8), alpha-ketoglutarate, urease (URH) and Bovine Serum Albumin (BSA).
2. The urea detection kit according to claim 1, wherein in the reagent 1: tris-succinate buffer (pH9.6) concentration 100-200mmol/L, alpha-ketoglutarate concentration 5.0-10.0mmol/L, glutamate dehydrogenase (GLDH) concentration 0.5-3.0KU/L, reduced coenzyme I (NADH) concentration 0.1-0.5mmol/L.
3. The urea detection kit of claim 1 or 2, wherein in the reagent 1, the optimal selective concentration of Tris-succinate buffer solution (pH 9.6) is 115mmol/L, the optimal selective concentration of alpha-ketoglutarate is 8mmol/L, the optimal selective concentration of glutamate dehydrogenase (GLDH) is 0.8KU/L, and the optimal selective concentration of reduced coenzyme I (NADH) is 0.35mmol/L.
4. The urea detection kit according to claim 1, wherein in the reagent 2: concentration of Tris buffer (ph 7.8): 40-150mmol/L, concentration of alpha-ketoglutaric acid: 5-15mmol/L, urease (URH) concentration: 20-50KU/L, bovine Serum Albumin (BSA) concentration: 0.8-1.5g/L.
5. The urea detection kit according to claim 4, wherein in the reagent 2: optimal selection concentration of Tris buffer (ph 7.8): 115mmol/L, optimum selective concentration of alpha-ketoglutaric acid: 8mmol/L, best selected concentration of Urease (URH): 40KU/L, bovine Serum Albumin (BSA) optimum selection concentration: 1g/L.
6. The urea detection kit of claim 1, wherein the glutamate dehydrogenase (GLDH) allosteric activator is one or both of Adenosine Diphosphate (ADP) and Guanine Dinucleotide Phosphate (GDP).
7. A urea detection kit according to claim 1 or 6, wherein the glutamate dehydrogenase (GLDH) allosteric activator is present in a concentration of 0.1mg/L to 5mg/L.
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CN112126674A (en) * | 2020-08-18 | 2020-12-25 | 兰州百源基因技术有限公司 | Urea detection kit, preparation method and use method thereof |
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Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
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CN112126674A (en) * | 2020-08-18 | 2020-12-25 | 兰州百源基因技术有限公司 | Urea detection kit, preparation method and use method thereof |
CN112126674B (en) * | 2020-08-18 | 2024-02-06 | 兰州百源基因技术有限公司 | Urea detection kit, preparation method and use method thereof |
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