CN1197974C - Digestive system tumor diagnosis kit - Google Patents
Digestive system tumor diagnosis kit Download PDFInfo
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- CN1197974C CN1197974C CN 01139260 CN01139260A CN1197974C CN 1197974 C CN1197974 C CN 1197974C CN 01139260 CN01139260 CN 01139260 CN 01139260 A CN01139260 A CN 01139260A CN 1197974 C CN1197974 C CN 1197974C
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- gpda
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- cancer
- reagent kit
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- 208000002699 Digestive System Neoplasms Diseases 0.000 title claims abstract description 5
- 238000003745 diagnosis Methods 0.000 title claims description 6
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 21
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 claims description 12
- YMAWOPBAYDPSLA-UHFFFAOYSA-N glycylglycine Chemical compound [NH3+]CC(=O)NCC([O-])=O YMAWOPBAYDPSLA-UHFFFAOYSA-N 0.000 claims description 8
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 6
- 238000012360 testing method Methods 0.000 claims description 6
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 claims description 6
- 108010008488 Glycylglycine Proteins 0.000 claims description 4
- 239000007983 Tris buffer Substances 0.000 claims description 4
- 229940043257 glycylglycine Drugs 0.000 claims description 4
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 claims description 3
- INEWUCPYEUEQTN-UHFFFAOYSA-N 3-(cyclohexylamino)-2-hydroxy-1-propanesulfonic acid Chemical compound OS(=O)(=O)CC(O)CNC1CCCCC1 INEWUCPYEUEQTN-UHFFFAOYSA-N 0.000 claims description 3
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 3
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 claims description 3
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 claims description 3
- 229940098773 bovine serum albumin Drugs 0.000 claims description 3
- 108010010043 glycylproline dipeptidyl aminopeptidase Proteins 0.000 abstract description 50
- 210000002966 serum Anatomy 0.000 abstract description 36
- 208000005718 Stomach Neoplasms Diseases 0.000 abstract description 14
- 230000000694 effects Effects 0.000 abstract description 13
- 201000007270 liver cancer Diseases 0.000 abstract description 9
- 208000014018 liver neoplasm Diseases 0.000 abstract description 9
- 206010017758 gastric cancer Diseases 0.000 abstract description 7
- 201000011549 stomach cancer Diseases 0.000 abstract description 7
- 239000000758 substrate Substances 0.000 abstract description 6
- 239000007788 liquid Substances 0.000 abstract description 4
- 238000012544 monitoring process Methods 0.000 abstract description 4
- XHLUWCXVWCBOPP-MERQFXBCSA-N (2s)-1-(2-aminoacetyl)-n-(4-nitrophenyl)pyrrolidine-2-carboxamide;4-methylbenzenesulfonic acid Chemical compound CC1=CC=C(S(O)(=O)=O)C=C1.NCC(=O)N1CCC[C@H]1C(=O)NC1=CC=C([N+]([O-])=O)C=C1 XHLUWCXVWCBOPP-MERQFXBCSA-N 0.000 abstract 1
- 230000000630 rising effect Effects 0.000 description 7
- 201000000498 stomach carcinoma Diseases 0.000 description 7
- 208000025865 Ulcer Diseases 0.000 description 6
- 231100000397 ulcer Toxicity 0.000 description 6
- 230000007423 decrease Effects 0.000 description 5
- 201000010099 disease Diseases 0.000 description 5
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 5
- 102100023635 Alpha-fetoprotein Human genes 0.000 description 3
- 208000007882 Gastritis Diseases 0.000 description 3
- 206010058467 Lung neoplasm malignant Diseases 0.000 description 3
- 108010026331 alpha-Fetoproteins Proteins 0.000 description 3
- 208000023652 chronic gastritis Diseases 0.000 description 3
- 201000005202 lung cancer Diseases 0.000 description 3
- 208000020816 lung neoplasm Diseases 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 210000000056 organ Anatomy 0.000 description 3
- 210000002784 stomach Anatomy 0.000 description 3
- 208000018522 Gastrointestinal disease Diseases 0.000 description 2
- 206010027457 Metastases to liver Diseases 0.000 description 2
- 206010028980 Neoplasm Diseases 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 201000004637 gastric cardia carcinoma Diseases 0.000 description 2
- 201000011066 hemangioma Diseases 0.000 description 2
- 230000002440 hepatic effect Effects 0.000 description 2
- 208000006454 hepatitis Diseases 0.000 description 2
- 230000003902 lesion Effects 0.000 description 2
- 210000001589 microsome Anatomy 0.000 description 2
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- 210000001519 tissue Anatomy 0.000 description 2
- 208000024893 Acute lymphoblastic leukemia Diseases 0.000 description 1
- 208000014697 Acute lymphocytic leukaemia Diseases 0.000 description 1
- 208000005623 Carcinogenesis Diseases 0.000 description 1
- 208000017897 Carcinoma of esophagus Diseases 0.000 description 1
- 206010008909 Chronic Hepatitis Diseases 0.000 description 1
- 108010035532 Collagen Proteins 0.000 description 1
- 102000008186 Collagen Human genes 0.000 description 1
- 101710173228 Glutathione hydrolase proenzyme Proteins 0.000 description 1
- 206010019728 Hepatitis alcoholic Diseases 0.000 description 1
- 206010019755 Hepatitis chronic active Diseases 0.000 description 1
- 206010023129 Jaundice cholestatic Diseases 0.000 description 1
- 206010027476 Metastases Diseases 0.000 description 1
- 208000015914 Non-Hodgkin lymphomas Diseases 0.000 description 1
- 201000005267 Obstructive Jaundice Diseases 0.000 description 1
- 206010030155 Oesophageal carcinoma Diseases 0.000 description 1
- 208000006664 Precursor Cell Lymphoblastic Leukemia-Lymphoma Diseases 0.000 description 1
- 108010077465 Tropocollagen Proteins 0.000 description 1
- 231100000354 acute hepatitis Toxicity 0.000 description 1
- 208000002353 alcoholic hepatitis Diseases 0.000 description 1
- 125000000539 amino acid group Chemical group 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 230000036952 cancer formation Effects 0.000 description 1
- 231100000504 carcinogenesis Toxicity 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 208000019425 cirrhosis of liver Diseases 0.000 description 1
- 229920001436 collagen Polymers 0.000 description 1
- 238000004737 colorimetric analysis Methods 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 208000010643 digestive system disease Diseases 0.000 description 1
- 201000005619 esophageal carcinoma Diseases 0.000 description 1
- 208000018685 gastrointestinal system disease Diseases 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- WGXUDTHMEITUBO-YFKPBYRVSA-N glutaurine Chemical compound OC(=O)[C@@H](N)CCC(=O)NCCS(O)(=O)=O WGXUDTHMEITUBO-YFKPBYRVSA-N 0.000 description 1
- 125000003630 glycyl group Chemical group [H]N([H])C([H])([H])C(*)=O 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 231100000283 hepatitis Toxicity 0.000 description 1
- 208000024557 hepatobiliary disease Diseases 0.000 description 1
- 230000003301 hydrolyzing effect Effects 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 210000005228 liver tissue Anatomy 0.000 description 1
- 206010025135 lupus erythematosus Diseases 0.000 description 1
- 208000025036 lymphosarcoma Diseases 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 239000000155 melt Substances 0.000 description 1
- 230000009401 metastasis Effects 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- ZRHANBBTXQZFSP-UHFFFAOYSA-M potassium;4-amino-3,5,6-trichloropyridine-2-carboxylate Chemical compound [K+].NC1=C(Cl)C(Cl)=NC(C([O-])=O)=C1Cl ZRHANBBTXQZFSP-UHFFFAOYSA-M 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
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- 206010039073 rheumatoid arthritis Diseases 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
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- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The present invention discloses a reagent kit for diagnosing tumor of the digestive system, which belongs to the technical field of biological reagents. The reagent kit is of a full liquid type with glycylprolyl p-nitroaniline p-toluene sulfonic acid (GPN) as a substrate for measuring the activity of glycylproline dipeptidyl aminopeptidase in blood serum. The reagent kit is used for diagnosing and monitoring liver cancer and stomach cancer. The reagent kit of the present invention has the advantages of good stability and very high clinical value.
Description
Technical field
The invention belongs to the biological reagent technical field.Be specifically related to a kind of digestive system tumor diagnosis kit.
Background technology
(Glycyl-proline-dipeptidylaminopeptidas is GPDA) from finding existing so far 30 years history for glycylproline dipeptidyl aminopeptidase.All there has been certain progress aspects such as fundamental research, clinical application and measuring method at GPDA.
As " Clinical Laboratory magazine " 1997 the 15th volume the 1st phase the 57th page of report: the rat of tentative liver cancer, the GPDA activity is approximately 2 times of healthy mouse in the serum, and the most GPDA specific activity of the homogenate of liver cancer tissue and microsome all significantly is lower than healthy mouse; And with the enchylema GPDA specific activity partly that liver cancer tissue is produced, be significantly higher than with normal liver tissue system taker.The rising of serum GPDA when liver cancer is described is to cause owing to the microsome endoenzyme outwards discharges to increase, and (γ-GT) rising is to increase cause machine-processed different owing to synthesizing with serum.
Patients with Gastric Cancer blood please obviously descend by GPDA, although decline mechanism is not clear, this phenomenon has been that many reports confirm.Full of interest is, with six kinds of different X-proline(Pro)-p-Nitroaniline (X-Pro-p-NA, the different amino-acid residue of X representative connection) makes substrate, measure the serum GPDA of normal people and Patients with Gastric Cancer, although the GPDA activity level that different substrates record differs fairly obvious, but every kind of active average of Patients with Gastric Cancer serum GPDA that substrate records separately all has only about half of normal people.In recent years some reports adopt the higher concentration substrate, have improved the active absolute value of GPDA, but do not change the multiple proportions relation of normal people and the active average of Patients with Gastric Cancer serum GPDA.
Other optimum gastrointestinal diseases, serum GPDA also can slightly descend.What decline by a big margin is stomach Collapse ulcer, is chronic gastritis and pyloric cap Collapse ulcer according to this.If " mean value-3 standard deviation " with the normal control group is boundary (X-3s), it is positive to be lower than this value, then the positive rate of stomach Collapse ulcer, chronic gastritis and pyloric cap Collapse ulcer is 18.4%, 6.2% and 0% according to this, far below the positive rate (72.5%) of cancer of the stomach.Serum GPDA is active during benign lesion descends, could be as the early signal of canceration, and Yet-have requires study.But, to the remarkable descender of serum GPDA, should be in conjunction with the possibility of clinical vigilant cancer of the stomach.Lung cancer, carcinoma of gastric cardia and cancer of esophagi human serum GPDA activity also have decline in various degree, and fall order according to this reduces.If when hepatic metastases appearred in cancer of the stomach and above-mentioned organ cancer, the GPDA activity can obviously raise.After the cancer of the stomach excision, serum GPDA has rise trend.
GPDA measures the main raw material in the reagent, be substrate---GPN (GPN), be mixed with after the liquid reagent extremely unstable, can only use the same day, otherwise must face with the preceding mixing that melts again in the following stored frozen of zero degrees celsius, once use, can not multigelation, bring very big inconvenience to practical application, there is carelessness very easily to influence the accuracy of measurement result slightly.Dry powder in the market and dry powder add the test kit of liquid, all this drawback of ubiquity.
Summary of the invention
Technical problem to be solved by this invention is to improve the stability of GPN under liquid state, makes the test kit of liquid-type.This test kit is made up of reagent 1 (R1) and reagent 2 (R2), and wherein reagent 1 is 0.5--3.7g% (W/V) Tutofusin tris (Tris), 0--0.1g% (W/V) NaN
3, 0.26--1.98g% (W/V) glycylglycine; Reagent 2 is 0--50mmol/L CAPSO (3-hexamethylene ammonia-2-hydroxyl-1-propane sulfonic acid sodium) or 0--0.5mmol/L trehalose, 5--50% (V/V) ethylene glycol or 30--70% (V/V) glycerol, 0--2g% bovine serum albumin, 5--40mmol/L GPN (glycyl L-prolyl-p-Nitroaniline-tosic acid) and 0--0.1g% (W/V) NaN
3
Test kit of the present invention both can be made continuous monitoring method, also can make 2 colorimetrys after adding stop buffer and use;
Work reagent absorbancy: R1 and R2 to mix A afterwards at 9: 1
405Nm≤1.0 (water school zero)
Precision: CV≤5%
Linearity range :≤638U/L
It is substrate that the present invention has succeeded in developing first with GPN, and stable full liquid-type GPDA measures test kit.
The clinical value that GPDA measures in the serum
The physiological function Yet-have of glycylproline dipeptidyl aminopeptidase (GPDA) is unclear, but because hydrolyzing N-end is the peptide bond of glycyl prolyl specifically, and exactly be rich in this constituent in the tropocollagen molecule, therefore infer in the degraded of collagen peptide, to play effect.The GPDA level is more constant in health adult's serum, and under some pathological state, can occur raising or reducing.Observe the variation of GPDA level in the serum, the diagnosis and the monitoring of disease had value.
The clinical meaning that GPDA measures in the serum mainly is:
1. to the diagnostic value of liver cancer and other hepatobiliary system diseases:
GPDA is active in the hepatocarcinoma patient serum significantly raises.Report was once arranged, and (healthy group is that except that a routine choledocholith, all the other 19 examples are liver cancer among 77.5 ± 17.1U/L) the 20 routine patients to serum GPDA＞200U/L.
The positive rate that GPDA raises among primary hepatocarcinoma (PHC) patients serum reaches about 83%, and secondary liver cancer can be up to 87.5%.The degree that two class patients serum GPDA raise is approaching, is about 1.5-2.5 times of healthy people; But also there is report secondary liver cancer patient to raise and reaches more than three times of normal healthy controls group.
PHC patients serum GPDA is active uncorrelated with alpha-fetoprotein (AFP), and subclinical phase patient PHC of several A FP feminine gender, and serum GPDA activity has been seen rising, illustrates that GPDA and AFP have good complementary action.But rising degree and the rising positive rate of subclinical phase PHC patients serum GPDA all significantly are lower than clinical phase patient, have reflected that serum GPDA level is relevant with the state of an illness of PHC.
Acute hepatitis, chronic active hepatitis, liver cirrhosis, obstructive jaundice etc., serum GPDA can have rising in various degree, and the rising amplitude is not as good as hepatocarcinoma patient.But when hepatitis gravis, alcoholic hepatitis, serum GPDA raises can be far more than hepatocarcinoma patient.
Hepatic hemangioma patient's serum GPDA activity is all within normal reference range.Therefore have the people to advocate, imaging diagnosis is the occupying lesion person, if serum GPDA raises, should get rid of hepatic hemangioma.
2. the activity level of cancer of the stomach and other digestive tract diseases, lung cancer patient serum GPDA:
GPDA is active in the Patients with Gastric Cancer serum obviously descends, and only reaches about half of healthy people's group; The about Patients with Gastric Cancer more than 70% of report is also arranged, and the X-3s that can drop to the normal healthy controls group is below horizontal.After the cancer of the stomach excision, serum GPDA has rise trend.
Carcinoma of gastric cardia, the esophageal carcinoma and lung cancer patient, serum GPDA can have decline in various degree.
When cancer of the stomach and above-mentioned other organ oncogenesis hepatic metastasess, serum GPDA can significantly raise.Dynamic observe the active variation of serum GPDA, help to guard the state of an illness of above-mentioned organ and liver metastasis whether.
Optimum gastrointestinal disease, serum GPDA also can slightly descend, and what decline by a big margin is stomach Collapse ulcer, is followed successively by chronic gastritis and pyloric cap Collapse ulcer.If with the positive boundary of the X-3s that is lower than the normal control group, then the positive rate of these three kinds of diseases is 18.4%, 6.2% and 0% according to this.
3. acute lymphoblastic leukemia, lymphosarcoma, Hokdkin disease and systemic lupus erythematous patient, and the course of disease is the rheumatoid arthritis patient more than 15 months, serum GPDA activity can significantly be lower than healthy people.
Embodiment
Embodiment 1
Reagent 1 (R1) contains:
0.155mol/L(1.88％W/V)Tris
0.065mol/L (0.86%W/V) glycylglycine
0.1％NaN
3
Reagent 2 (R2) contains:
33% (V/V) ethylene glycol
20mmol/L?CAPSO
1% bovine serum albumin
0.05％NaN
3
30mmol/L?GPN
Embodiment 2
Reagent 1 (R1) contains:
0.155mol/L(1.88％W/V)Tris
0.065mol/L (0.86%W/V) glycylglycine
0.1％NaN
3
Reagent 2 (R2) contains:
33% (V/V) ethylene glycol
0.25mmol/L trehalose
0.055％NaN
3
30mmol/L?GPN
Embodiment 3
Measure the GPDA activity methods:
Wavelength (main/time) 405nm/600nm or 410nm/600nm
Optical path 1cm
37 ℃ of temperature
Sample 13 μ l R1 234 μ l R2 26 μ l
Lag time 30 seconds
Monitoring time 60 seconds
Input constant 405nm/600nm:2126; 410hm/600nm:2457
Claims (1)
1, a kind of digestive system tumor diagnosis kit is characterized in that this test kit is made up of reagent 1 (R1) and reagent 2 (R2), and wherein reagent 1 is 0.5--3.7g% (W/V) Tutofusin tris (Tris), 0--0.1g% (W/V) NaN
3, 0.26--1.98g% (W/V) glycylglycine; Reagent 2 is 0--50mmol/L CAPSO (3-hexamethylene ammonia-2-hydroxyl-1-propane sulfonic acid sodium) or 0--0.5mmol/L trehalose, 5--50% (V/V) ethylene glycol or 30--70% (V/V) glycerol, 0--2g% bovine serum albumin, 5--40mmol/L GPN (glycyl-L-prolyl-p-Nitroaniline-tosic acid) and 0--0.1g% (W/V) NaN
3
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 01139260 CN1197974C (en) | 2001-12-27 | 2001-12-27 | Digestive system tumor diagnosis kit |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 01139260 CN1197974C (en) | 2001-12-27 | 2001-12-27 | Digestive system tumor diagnosis kit |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1428436A CN1428436A (en) | 2003-07-09 |
| CN1197974C true CN1197974C (en) | 2005-04-20 |
Family
ID=4675182
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 01139260 Expired - Lifetime CN1197974C (en) | 2001-12-27 | 2001-12-27 | Digestive system tumor diagnosis kit |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1197974C (en) |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100353166C (en) * | 2004-08-30 | 2007-12-05 | 北京源德生物医学工程有限公司 | Enzyme tag stabilizing agent |
| CN104880423A (en) * | 2015-05-08 | 2015-09-02 | 浙江蓝森生物科技有限公司 | Method, reagent and kit for quantitatively determining activity of glycylproline dipeptidyl aminopeptidase in human serum |
| CN105510572A (en) * | 2015-12-22 | 2016-04-20 | 山东博科生物产业有限公司 | Detection kit for glycylproline dipeptidyl aminopeptidase |
| CN105861632A (en) * | 2016-04-28 | 2016-08-17 | 安徽伊普诺康生物技术股份有限公司 | Kit for determining glycylproline dipeptide aminopeptidase |
-
2001
- 2001-12-27 CN CN 01139260 patent/CN1197974C/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| CN1428436A (en) | 2003-07-09 |
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