CN1635379A - Serum copper, iron and zinc rapid sensitive detection kit and preparing method thereof - Google Patents
Serum copper, iron and zinc rapid sensitive detection kit and preparing method thereof Download PDFInfo
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- CN1635379A CN1635379A CN 200410096340 CN200410096340A CN1635379A CN 1635379 A CN1635379 A CN 1635379A CN 200410096340 CN200410096340 CN 200410096340 CN 200410096340 A CN200410096340 A CN 200410096340A CN 1635379 A CN1635379 A CN 1635379A
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- pyridylazo
- bromo
- phenol
- colour developing
- serum
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- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 title claims abstract description 74
- 210000002966 serum Anatomy 0.000 title claims abstract description 53
- 229910052742 iron Inorganic materials 0.000 title claims abstract description 37
- 239000011701 zinc Substances 0.000 title claims abstract description 36
- 239000010949 copper Substances 0.000 title claims abstract description 34
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 title claims abstract description 32
- 229910052725 zinc Inorganic materials 0.000 title claims abstract description 32
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 title claims abstract description 30
- 229910052802 copper Inorganic materials 0.000 title claims abstract description 30
- 238000011896 sensitive detection Methods 0.000 title claims description 21
- 238000000034 method Methods 0.000 title abstract description 19
- 239000007788 liquid Substances 0.000 claims abstract description 50
- 239000003795 chemical substances by application Substances 0.000 claims abstract description 22
- 238000002360 preparation method Methods 0.000 claims description 24
- 239000011550 stock solution Substances 0.000 claims description 24
- 239000004094 surface-active agent Substances 0.000 claims description 21
- 239000003153 chemical reaction reagent Substances 0.000 claims description 19
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 18
- 238000005303 weighing Methods 0.000 claims description 18
- -1 (2) pyridylazo Chemical class 0.000 claims description 16
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 16
- 238000012216 screening Methods 0.000 claims description 16
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims description 15
- 229920001214 Polysorbate 60 Polymers 0.000 claims description 14
- 239000007864 aqueous solution Substances 0.000 claims description 14
- BHZOKUMUHVTPBX-UHFFFAOYSA-M sodium acetic acid acetate Chemical compound [Na+].CC(O)=O.CC([O-])=O BHZOKUMUHVTPBX-UHFFFAOYSA-M 0.000 claims description 12
- UMGDCJDMYOKAJW-UHFFFAOYSA-N thiourea Chemical compound NC(N)=S UMGDCJDMYOKAJW-UHFFFAOYSA-N 0.000 claims description 12
- 229920003171 Poly (ethylene oxide) Polymers 0.000 claims description 10
- CWERGRDVMFNCDR-UHFFFAOYSA-N thioglycolic acid Chemical compound OC(=O)CS CWERGRDVMFNCDR-UHFFFAOYSA-N 0.000 claims description 10
- 229920004890 Triton X-100 Polymers 0.000 claims description 8
- 239000013504 Triton X-100 Substances 0.000 claims description 8
- LWZFANDGMFTDAV-BURFUSLBSA-N [(2r)-2-[(2r,3r,4s)-3,4-dihydroxyoxolan-2-yl]-2-hydroxyethyl] dodecanoate Chemical compound CCCCCCCCCCCC(=O)OC[C@@H](O)[C@H]1OC[C@H](O)[C@H]1O LWZFANDGMFTDAV-BURFUSLBSA-N 0.000 claims description 8
- LNIGBQNXJNNICD-UHFFFAOYSA-N 2-[(5-bromopyridin-2-yl)diazenyl]-5-(dimethylamino)phenol Chemical compound OC1=CC(N(C)C)=CC=C1N=NC1=CC=C(Br)C=N1 LNIGBQNXJNNICD-UHFFFAOYSA-N 0.000 claims description 7
- ZVZFHCZCIBYFMZ-UHFFFAOYSA-N 6-methylheptoxybenzene Chemical compound CC(C)CCCCCOC1=CC=CC=C1 ZVZFHCZCIBYFMZ-UHFFFAOYSA-N 0.000 claims description 7
- HOBSGKWFTBDQCH-UHFFFAOYSA-N phenol;sodium Chemical compound [Na].[Na].OC1=CC=CC=C1 HOBSGKWFTBDQCH-UHFFFAOYSA-N 0.000 claims description 7
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims description 7
- WRIDQFICGBMAFQ-UHFFFAOYSA-N (E)-8-Octadecenoic acid Natural products CCCCCCCCCC=CCCCCCCC(O)=O WRIDQFICGBMAFQ-UHFFFAOYSA-N 0.000 claims description 6
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 claims description 6
- LQJBNNIYVWPHFW-UHFFFAOYSA-N 20:1omega9c fatty acid Natural products CCCCCCCCCCC=CCCCCCCCC(O)=O LQJBNNIYVWPHFW-UHFFFAOYSA-N 0.000 claims description 6
- QSBYPNXLFMSGKH-UHFFFAOYSA-N 9-Heptadecensaeure Natural products CCCCCCCC=CCCCCCCCC(O)=O QSBYPNXLFMSGKH-UHFFFAOYSA-N 0.000 claims description 6
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 6
- 239000005642 Oleic acid Substances 0.000 claims description 6
- ZQPPMHVWECSIRJ-UHFFFAOYSA-N Oleic acid Natural products CCCCCCCCC=CCCCCCCCC(O)=O ZQPPMHVWECSIRJ-UHFFFAOYSA-N 0.000 claims description 6
- QXJSBBXBKPUZAA-UHFFFAOYSA-N isooleic acid Natural products CCCCCCCC=CCCCCCCCCC(O)=O QXJSBBXBKPUZAA-UHFFFAOYSA-N 0.000 claims description 6
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 claims description 6
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 claims description 6
- 229920000036 polyvinylpyrrolidone Polymers 0.000 claims description 6
- 239000001267 polyvinylpyrrolidone Substances 0.000 claims description 6
- PUZPDOWCWNUUKD-UHFFFAOYSA-M sodium fluoride Chemical compound [F-].[Na+] PUZPDOWCWNUUKD-UHFFFAOYSA-M 0.000 claims description 6
- 239000007787 solid Substances 0.000 claims description 6
- 239000000243 solution Substances 0.000 claims description 6
- WAVOOWVINKGEHS-UHFFFAOYSA-N 3-(diethylamino)phenol Chemical compound CCN(CC)C1=CC=CC(O)=C1 WAVOOWVINKGEHS-UHFFFAOYSA-N 0.000 claims description 4
- GDEBSAWXIHEMNF-UHFFFAOYSA-O cupferron Chemical compound [NH4+].O=NN([O-])C1=CC=CC=C1 GDEBSAWXIHEMNF-UHFFFAOYSA-O 0.000 claims description 4
- USHAGKDGDHPEEY-UHFFFAOYSA-L potassium persulfate Chemical compound [K+].[K+].[O-]S(=O)(=O)OOS([O-])(=O)=O USHAGKDGDHPEEY-UHFFFAOYSA-L 0.000 claims description 4
- GCLGEJMYGQKIIW-UHFFFAOYSA-H sodium hexametaphosphate Chemical compound [Na]OP1(=O)OP(=O)(O[Na])OP(=O)(O[Na])OP(=O)(O[Na])OP(=O)(O[Na])OP(=O)(O[Na])O1 GCLGEJMYGQKIIW-UHFFFAOYSA-H 0.000 claims description 4
- 235000019982 sodium hexametaphosphate Nutrition 0.000 claims description 4
- 239000001577 tetrasodium phosphonato phosphate Substances 0.000 claims description 4
- 235000017858 Laurus nobilis Nutrition 0.000 claims description 3
- 235000005212 Terminalia tomentosa Nutrition 0.000 claims description 3
- 235000013024 sodium fluoride Nutrition 0.000 claims description 3
- 239000011775 sodium fluoride Substances 0.000 claims description 3
- 235000015165 citric acid Nutrition 0.000 claims description 2
- 244000125380 Terminalia tomentosa Species 0.000 claims 1
- 150000001875 compounds Chemical class 0.000 abstract description 6
- 102000004169 proteins and genes Human genes 0.000 abstract description 3
- 108090000623 proteins and genes Proteins 0.000 abstract description 3
- 238000012360 testing method Methods 0.000 abstract description 2
- 150000002894 organic compounds Chemical class 0.000 abstract 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 24
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 12
- 239000012153 distilled water Substances 0.000 description 10
- 238000001514 detection method Methods 0.000 description 8
- 238000002835 absorbance Methods 0.000 description 7
- 238000006243 chemical reaction Methods 0.000 description 7
- 238000003556 assay Methods 0.000 description 6
- 238000006757 chemical reactions by type Methods 0.000 description 6
- 230000003287 optical effect Effects 0.000 description 6
- 238000004321 preservation Methods 0.000 description 6
- CWYNVVGOOAEACU-UHFFFAOYSA-N Fe2+ Chemical compound [Fe+2] CWYNVVGOOAEACU-UHFFFAOYSA-N 0.000 description 4
- GLUUGHFHXGJENI-UHFFFAOYSA-N Piperazine Chemical compound C1CNCCN1 GLUUGHFHXGJENI-UHFFFAOYSA-N 0.000 description 4
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 description 4
- 230000002829 reductive effect Effects 0.000 description 4
- 208000007502 anemia Diseases 0.000 description 3
- 238000004737 colorimetric analysis Methods 0.000 description 3
- 230000035945 sensitivity Effects 0.000 description 3
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
- 244000147568 Laurus nobilis Species 0.000 description 2
- 208000031845 Pernicious anaemia Diseases 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 238000013459 approach Methods 0.000 description 2
- 238000003321 atomic absorption spectrophotometry Methods 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- 239000008367 deionised water Substances 0.000 description 2
- 229910021641 deionized water Inorganic materials 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 description 2
- 238000002798 spectrophotometry method Methods 0.000 description 2
- VTLYFUHAOXGGBS-UHFFFAOYSA-N Fe3+ Chemical compound [Fe+3] VTLYFUHAOXGGBS-UHFFFAOYSA-N 0.000 description 1
- 206010018910 Haemolysis Diseases 0.000 description 1
- 208000002972 Hepatolenticular Degeneration Diseases 0.000 description 1
- 208000017604 Hodgkin disease Diseases 0.000 description 1
- 208000010747 Hodgkins lymphoma Diseases 0.000 description 1
- WTDHULULXKLSOZ-UHFFFAOYSA-N Hydroxylamine hydrochloride Chemical compound Cl.ON WTDHULULXKLSOZ-UHFFFAOYSA-N 0.000 description 1
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 description 1
- 208000012641 Pigmentation disease Diseases 0.000 description 1
- 244000061458 Solanum melongena Species 0.000 description 1
- 102000004338 Transferrin Human genes 0.000 description 1
- 108090000901 Transferrin Proteins 0.000 description 1
- PTFCDOFLOPIGGS-UHFFFAOYSA-N Zinc dication Chemical compound [Zn+2] PTFCDOFLOPIGGS-UHFFFAOYSA-N 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 229960005070 ascorbic acid Drugs 0.000 description 1
- 235000010323 ascorbic acid Nutrition 0.000 description 1
- 239000011668 ascorbic acid Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 230000032823 cell division Effects 0.000 description 1
- 239000013522 chelant Substances 0.000 description 1
- RNFNDJAIBTYOQL-UHFFFAOYSA-N chloral hydrate Chemical compound OC(O)C(Cl)(Cl)Cl RNFNDJAIBTYOQL-UHFFFAOYSA-N 0.000 description 1
- 229960002327 chloral hydrate Drugs 0.000 description 1
- 230000001684 chronic effect Effects 0.000 description 1
- 238000007398 colorimetric assay Methods 0.000 description 1
- 230000000536 complexating effect Effects 0.000 description 1
- 230000008588 hemolysis Effects 0.000 description 1
- 208000007475 hemolytic anemia Diseases 0.000 description 1
- 230000008676 import Effects 0.000 description 1
- 235000021125 infant nutrition Nutrition 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 230000002452 interceptive effect Effects 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- JEIPFZHSYJVQDO-UHFFFAOYSA-N iron(III) oxide Inorganic materials O=[Fe]O[Fe]=O JEIPFZHSYJVQDO-UHFFFAOYSA-N 0.000 description 1
- 208000032839 leukemia Diseases 0.000 description 1
- 229910017604 nitric acid Inorganic materials 0.000 description 1
- 230000004962 physiological condition Effects 0.000 description 1
- 231100000572 poisoning Toxicity 0.000 description 1
- 230000000607 poisoning effect Effects 0.000 description 1
- NNFCIKHAZHQZJG-UHFFFAOYSA-N potassium cyanide Chemical compound [K+].N#[C-] NNFCIKHAZHQZJG-UHFFFAOYSA-N 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 208000008128 pulmonary tuberculosis Diseases 0.000 description 1
- 239000000376 reactant Substances 0.000 description 1
- 208000011580 syndromic disease Diseases 0.000 description 1
- 238000004448 titration Methods 0.000 description 1
- 239000011573 trace mineral Substances 0.000 description 1
- 235000013619 trace mineral Nutrition 0.000 description 1
- 239000012581 transferrin Substances 0.000 description 1
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- Investigating Or Analysing Biological Materials (AREA)
- Investigating Or Analyzing Non-Biological Materials By The Use Of Chemical Means (AREA)
Abstract
This invention discloses a test agent box to measure the contents of copper, iron and zinc in blood serum and provides the process method of the agent box. The agent box in this invention dissociates the protein with copper, iron and zinc in blood serum in the buffer liquid system with the surface activity agent, which is compound with the pyridylazo organic compound to make color. The color complex compound has the longest wavelength at 550-600nm.
Description
Technical field
The present invention relates to be respectively applied for detection Copper in Serum, iron, the rapid sensitive detection kit of zinc content and the preparation method of each kit thereof.
Background technology
Copper, iron, zinc are the trace elements of needed by human.
Serum copper increases and sees leukaemia, Hodgkin's disease, pigmentation disease, pulmonary tuberculosis, pernicious anaemia, malignant tumour etc.The serum copper reduction sees baby's anaemia, the bad Menkes curly hair of infant nutrition syndrome, hepatolenticular degeneration etc.The method of measuring serum copper is a lot, first-selected clinically at present method is that this method is adopted in atomic absorption spectrophotometry, external most of laboratory, but this method instrument costliness, in China clinical labororatory, be difficult to popularize, what extensively test clinically at present is spectrophotometric method, the spectrophotometric method first method is to adopt bicyclohexanone acyl dihydrazone (abbreviation cupferron), and this method sensitivity is low, and (ε is 1.6 * 10
4Lmol
-1Cm
-1), the serum consumption is big, and needs deproteinized, and the running time is long, numerous; Second method is to adopt 2,9-dimethyl 4, and 7-diphenyl-1, the 10-ferrosin, this method sensitivity is not high, and (ε is 1.4 * 10
4Lmol
-1Cm
-1), the reagent price is expensive, needs import.
Zinc is the accessory factor of many metalloenzyme.Because synthetic DNA, the essential many metalloenzyme of RNA all need zinc, so it is that cell division institute is requisite, is so in the high-speed rapid growth phase of life especially, as baby, teenager, pregnant woman.Serum zinc has the laudatory title of " flower of life ".Serum zinc raises and is common in zinc poisoning.At present, the assay method of zinc is more, but for a long time, still lacks the detection method that can promote clinically.Complex titration method amount of samples big (serum 2ml), during operating cost, specificity is low; Fluorophotometric method instrument price is expensive, is difficult to popularize; Also there are shortcomings such as operation is numerous, cost height in atomic absorption spectrophotometry.Currently reported spectrophotometric colourimetry all need be introduced the ascorbic acid reduction, and hypertoxic potassium cyanide is sheltered interfering ion, demasks with chloral hydrate again.Because of its reagent type is many, operation steps is numerous and require high and be difficult to promote.
Serum levels of iron reduces, and is common in the iron that physiological iron requirement increases, various chronic blood loss causes and loses too much and the iron insufficiency of intake, as hypoferric anemia, acute infection, malignant tumour etc.Serum levels of iron increases, and is common in oxyhepatitis, pernicious anaemia, alpastic anemia, hemolytic anemia etc.The serum levels of iron horizontal instability is subject to the influence of feed situation and other physiological conditions.At present, the assay method of iron commonly used has ferrous piperazine colourimetry, utilize serum levels of iron and transferrin to be combined into compound, iron dissociates out from compound in acid medium, is reduced agent again and is reduced into ferrous iron, and generate the aubergine compound with ferrous piperazine, at wavelength 562nm place one absorption peak is arranged, compare with the titer of same processing, can try to achieve the content of serum levels of iron, this method costs an arm and a leg, step is numerous and diverse; Utilize the duplex pyridine colorimetric method in addition, its be utilize under acid condition, make iron from the protein bound state dissociate out.Make as reductive agent with oxammonium hydrochloride that ferric iron is reduced into ferrous iron in the serum, the latter and the reaction of duplex pyridine developer generate red chelate, at 520nm place colorimetric assay.This law is easy fast, but poor sensitivity, disturbing factor is more, and the most serious with the irony pollution especially, haemolysis also can influence the result.
Summary of the invention
Technical matters to be solved by this invention provides a kind of kit that is used to measure the easy and simple to handle quick, highly sensitive of Copper in Serum, iron, zinc content and does not need deproteinized, and for this reason, the present invention also will provide a kind of preparation method of this kit.
The principle of kit of the present invention is: in the buffer system in the presence of the surfactant, dissociate with protein bound copper, iron, zinc in the serum, develop the color with the complexing of pyridylazo type organic, the colour developing complex compound has maximum absorption wavelength at wavelength 550-600nm place, reactant liquor is directly proportional with copper, iron, zinc ion concentration within the specific limits in the absorbance at wavelength 550-600nm place, is suitable for manual mensuration and automatic the analysis.
For solving the problems of the technologies described above, the technical solution used in the present invention is:
1, be used to detect the rapid sensitive detection kit of content of Cu in serum, it is made up of following reagent:
(1) oxygenant: concentration is hydrogen peroxide or the potassium persulfate of 30g/L;
(2) pyridylazo class colour developing liquid: solid content is by 20-100mg pyridylazo type organic in every liter of colour developing liquid, 5-20g surfactant and 20-100g screening agent are formed, liquid by pH=3.5-5.0 acetic acid-sodium acetate aqueous solution and absolute ethyl alcohol in 199: 1-19: 1 ratio is formed, described surfactant is that described surfactant is a polyoxyethylene laurel ether, polyoxyethylene iso-octyl phenyl ether, Triton X-100, polyoxyethylene sorbitan lauric acid monoester, polyoxyethylene sorbitan oleic acid monoester or polyvinylpyrrolidone, described screening agent are citric acid, sodium fluoride or sodium hexametaphosphate.
(3) copper titer: concentration is 15.74-31.48 μ mol/L.
The preparation method of described pyridylazo class colour developing liquid is: every liter of colour developing liquid takes by weighing pyridylazo type organic 100-500mg, be made into the 100ml stock solution with absolute ethyl alcohol, get stock solution 5-20ml, add the surfactant of 5-20g, be diluted to 1000ml with acetic acid-sodium acetate solution of pH=3.5-5.0.
Described pyridylazo type organic is 2-(5-bromo-2-pyridylazo)-5-(diethylamino phenol), 2-(3,5-two bromo-2-pyridylazos)-and 5-diethylamino phenol, 2-(5-bromo-2-pyridylazo)-5-dimethylamino phenol, 2-(3,5-two bromo-2-pyridylazos)-5-dimethylamino phenol or 2-(5-nitro-2-pyridylazo)-5-[N-n-pro-pyl-N-(3-sulfonic acid propyl group) amino] the phenol disodium.
2, be used to detect the rapid sensitive detection kit of Zn in serum content, it is characterized in that: it is made up of following reagent:
(1) pyridylazo class colour developing liquid: solid content is by 20-100mg pyridylazo type organic in every liter of colour developing liquid, 5-20g surfactant and 20-100g screening agent are formed, liquid by the Tri(Hydroxymethyl) Amino Methane Hydrochloride aqueous solution of pH=8.0-10 and absolute ethyl alcohol in 199: 1-19: 1 ratio is formed, described surfactant is a polyoxyethylene iso-octyl phenyl ether, Triton X-100, polyoxyethylene sorbitan lauric acid monoester, polyoxyethylene sorbitan oleic acid monoester or polyvinylpyrrolidone, described screening agent are cupferron, citric acid, mercaptoacetic acid, thiocarbamide or sodium hexametaphosphate.
(2) zinc titer: concentration is 15.3-30.6 μ mol/L.
The preparation method of described pyridylazo class colour developing liquid is: every liter of colour developing liquid takes by weighing the screening agent of pyridylazo type organic 100-500mg and 20-100g, be made into the 100ml stock solution with absolute ethyl alcohol, get stock solution 5-50ml, the surfactant that adds 5-20g is diluted to 1000ml with the Tri(Hydroxymethyl) Amino Methane Hydrochloride aqueous solution of pH=8.0-10.0.
Described pyridylazo type organic is 2-(5-bromo-2-pyridylazo)-5-(diethylamino phenol), 2-(3,5-two bromo-2-pyridylazos)-5-diethylamino phenol, 2-(5-bromo-2-pyridylazo)-5-dimethylamino phenol, 2-(3,5-two bromo-2-pyridylazos)-5-dimethylamino phenol (be called for short 3,5-diBr-DMPAP) or 2-(5-nitro-2-pyridylazo)-5-[N-n-pro-pyl-N-(3-sulfonic acid propyl group) amino] the phenol disodium.
3, be used for detecting the rapid sensitive detection kit of serum iron content, it is made up of following reagent:
(1) pyridylazo class colour developing liquid: solid content is made up of 20-100mg pyridylazo type organic, 5-20g surfactant and 20-100g screening agent in every liter of colour developing liquid, liquid by pH=3.5-5.0 acetic acid-sodium acetate aqueous solution and absolute ethyl alcohol in 199: 1-19: 1 ratio is formed, described surfactant is polyoxyethylene iso-octyl phenyl ether, Triton X-100, polyoxyethylene sorbitan lauric acid monoester, polyoxyethylene sorbitan oleic acid monoester or polyvinylpyrrolidone, and described screening agent is mercaptoacetic acid or thiocarbamide.
(2) iron titer: concentration is 17.9-35.8 μ mol/L.
The preparation method of described pyridylazo class colour developing liquid is: take by weighing pyridylazo type organic 100-500mg and 20-100g screening agent, be made into the 100ml stock solution with absolute ethyl alcohol, get stock solution 5-50ml, the surfactant that adds 5-20g is diluted to 1000ml with acetic acid-sodium acetate solution of pH=3.5-5.0.
Described pyridylazo type organic is 2-(5-bromo-2-pyridylazo)-5-(diethylamino phenol), 2-(3,5-two bromo-2-pyridylazos)-and 5-diethylamino phenol, 2-(5-bromo-2-pyridylazo)-5-dimethylamino phenol, 2-(3,5-two bromo-2-pyridylazos)-5-dimethylamino phenol or 2-(5-nitro-2-pyridylazo)-5-[N-n-pro-pyl-N-(3-sulfonic acid propyl group) amino] the phenol disodium.
The beneficial effect that the present invention can reach is: the serum consumption is few, the single Wen Ding of reagent, and needn't deproteinized operation fast and convenient, accuracy is good, and is highly sensitive.
Embodiment
Embodiment one
A kind of serum copper rapid sensitive detection kit, this kit is made up of following material:
1 bottle in oxygenant (hydrogen peroxide of 30g/L)
Colour developing liquid (pyridylazo class developer) 1 bottle
1 bottle of copper titer (concentration is 31.48 μ mol/L)
The preparation method of this kit realizes as follows:
(1) hydrogen peroxide of 30g/L is the conventional method configuration;
(2) preparation method of pyridylazo class colour developing liquid:
A, take by weighing 2-(5-bromo-2-pyridylazo)-5-(diethylamino phenol) 200mg,
B, the reagent among a is made into the stock solution of 100ml with absolute ethyl alcohol,
C, the stock solution 5ml that gets b add 10g polyoxyethylene iso-octyl phenyl ether and 20g citric acid, are diluted to 1000ml with acetic acid-sodium acetate aqueous solution of pH4.0;
(3) copper titer: accurately take by weighing the metallic copper (99.9%) of 100.0mg, the aqueous acetic acid dissolving back with an amount of 50% is diluted to 1000ml with distilled water, and dilute with water is the copper titer of 50 times 31.48 μ mol/L again.
Detection method:
1. manual analytic approach: serum (U pipe), copper titer (S pipe), deionized water (B pipe) are respectively got 0.20ml, in each pipe, add 3% hydrogen peroxide 0.05ml then respectively, left standstill 1 minute, add 2.5ml pyridylazo class colour developing liquid more respectively, mixing, put 37 ℃ of water bath heat preservations 5 minutes, and can guarantee that serum copper disengaged fully.On spectrophotometer,, read the absorbance (A) of each pipe then at the 550-600nm place with the blank pipe zeroing of 1cm optical path cuvette.
2. automatic analyzer analytical parameters: reaction type: end-point method; Wavelength: 600nm; Temperature of reaction: 37 ℃; 3% hydrogen peroxide 5 μ l; Pyridylazo class colour developing liquid: 250 μ l; Sample: 30 μ l; Reading duration: 180 seconds.
The content of serum copper (μ mol/L)=31.48 * Au/As
Embodiment two
A kind of serum copper rapid sensitive detection kit, this kit is made up of following material:
1 bottle in oxygenant (potassium persulfate of 30g/L)
Colour developing liquid (pyridylazo class developer) 1 bottle
1 bottle of copper titer (concentration is 15.74 μ mol/L)
The preparation method of this kit realizes as follows:
(1) potassium persulfate of 30g/L is the conventional method configuration
(2) preparation method of pyridylazo class developer:
A, take by weighing 2-(3,5-two bromo-2-pyridylazos)-5-diethylamino phenol 100mg,
B, the reagent among a is made into the stock solution of 100ml with absolute ethyl alcohol,
C, the stock solution 50ml that gets b add 20g polyoxyethylene laurel ether and 50g sodium fluoride, are diluted to 1000ml with acetic acid-sodium acetate aqueous solution of PH3.8.
(3) copper titer: accurately take by weighing the metallic copper (99.9%) of 100.0mg, the aqueous acetic acid dissolving back with an amount of 50% is diluted to 1000ml with distilled water, and dilute with water is the copper titer of 100 times 15.74 μ mol/L again.
Detection method:
1. manual analytic approach: serum (U pipe), copper titer (S pipe), deionized water (B pipe) are respectively got 0.20ml, in each pipe, add 3% hydrogen peroxide 0.05ml then respectively, left standstill 1 minute, add 2.5ml pyridylazo class colour developing liquid more respectively, mixing, put 37 ℃ of water bath heat preservations 5 minutes, and can guarantee that serum copper disengaged fully.On spectrophotometer,, read the absorbance (A) of each pipe then at the 550-600nm place with the blank pipe zeroing of 1cm optical path cuvette.
2. automatic analyzer analytical parameters: reaction type: end-point method; Wavelength: 600nm; Temperature of reaction: 37 ℃; 3% hydrogen peroxide 5 μ l; Pyridylazo class colour developing liquid: 250 μ l; Sample: 30 μ l; Reading duration: 180 seconds.
The content of serum copper (μ mol/L)=15.74 * Au/As
Embodiment three
A kind of serum zinc rapid sensitive detection kit, this kit is made up of following reagent:
Colour developing liquid (pyridylazo class developer) 1 bottle
1 bottle of zinc titer (concentration is 15.3 μ mol/L)
The preparation method of this kit realizes as follows:
(1) preparation method of pyridylazo class colour developing liquid:
A, take by weighing 2-(5-bromo-2-pyridylazo)-5-dimethylamino phenol 300mg,
B, the reagent among a is made into the stock solution of 100ml with absolute ethyl alcohol,
C, the stock solution 30ml that gets b add the cupferron of 15g polyoxyethylene sorbitan lauric acid monoester and 20g, are diluted to 1000ml with the Tri(Hydroxymethyl) Amino Methane Hydrochloride aqueous solution of pH8.0.
(2) zinc titer: accurately take by weighing the metallic zinc (99.9%) of 100.0mg, the salpeter solution dissolving back with an amount of 50% is diluted to 1000ml with distilled water, and dilute with water is the zinc titer of 15.30 μ mol/L again.
Detection method:
1. manual assay pipe (U), standard pipe (S), blank pipe (B) add serum, zinc titer, each 0.3ml of distilled water respectively, every pipe adds pyridylazo class colour developing liquid 2.50ml, mixing, put 37 ℃ of water bath heat preservations 5 minutes, return to zero with the blank pipe, with 1cm optical path cuvette, mensuration is respectively managed absorbance A at 550--600nm wavelength place.
2. automatic analyzer analytical parameters reaction type: end-point method; Wavelength: 600nm; Temperature of reaction: 37 ℃; Pyridylazo class colour developing liquid: 250 μ l; Sample: 30 μ l; Reading duration: 180 seconds.
The content of serum zinc (μ mol/L)=15.30 * Au/As
Embodiment four
A kind of serum zinc rapid sensitive detection kit, this kit is made up of following reagent:
Colour developing liquid (pyridylazo class developer) 1 bottle
1 bottle of zinc titer (concentration is 30.6 μ mol/L)
The preparation method of this kit realizes as follows:
(1) preparation method of pyridylazo class developer:
A, take by weighing 2-(5-nitro-2-pyridylazo)-5-[N-n-pro-pyl-N-(3-sulfonic acid propyl group) amino] phenol disodium 150mg,
B, the reagent among a is made into the stock solution of 100ml with absolute ethyl alcohol,
C, the stock solution 15ml that gets b add the thiocarbamide of 15g Triton X-100 and 80g, are diluted to 1000ml with the Tri(Hydroxymethyl) Amino Methane Hydrochloride aqueous solution of PH8.0.
(2) zinc titer: accurately take by weighing the metallic zinc (99.9%) of 100.0mg, the aqueous acetic acid dissolving back with an amount of 50% is diluted to 1000ml with distilled water, and dilute with water is the zinc titer of 50 times 30.60 μ mol/L again.
Detection method:
1. manual assay pipe (U), standard pipe (S), blank pipe (B) add serum, iron titer, each 0.3ml of distilled water respectively, every pipe adds pyridylazo class colour developing liquid 2.50ml, mixing, put 37 ℃ of water bath heat preservations 5 minutes, return to zero with the blank pipe, with 1cm optical path cuvette, mensuration is respectively managed absorbance A at 550--600nm wavelength place.
2. automatic analyzer analytical parameters reaction type: end-point method; Wavelength: 600nm; Temperature of reaction: 37 ℃; Pyridylazo class colour developing liquid: 250 μ l; Sample: 30 μ l; Reading duration: 180 seconds.
The content of serum zinc (μ mol/L)=30.60 * Au/As
Embodiment five
A kind of serum levels of iron rapid sensitive detection kit, this kit is made up of following reagent:
Colour developing liquid (pyridylazo class developer) 1 bottle
1 bottle of iron titer (concentration is 17.9 μ mol/L)
The preparation method of this kit realizes as follows:
(2) preparation method of pyridylazo class colour developing liquid:
A, take by weighing 2-(3,5-two bromo-2-pyridylazos)-5-dimethylamino phenol 300mg,
B, the reagent among a is made into the stock solution of 100ml with absolute ethyl alcohol,
C, the stock solution 30ml that gets b add the mercaptoacetic acid of 15g polyoxyethylene sorbitan lauric acid monoester and 20g, are diluted to 1000ml with the acetic acid sodium acetate aqueous solution of pH3.5.
(3) iron titer (concentration is 17.9 μ mol/L): accurately take by weighing the metallic iron (99.9%) of 100.0mg, with being diluted to 1000ml with distilled water behind an amount of nitric acid dissolve, dilute with water is that concentration is 17.9 μ mol/L titers again.
Detection method:
1. manual assay pipe (U), standard pipe (S), blank pipe (B) add serum, iron titer, each 0.3ml of distilled water respectively, every pipe adds pyridylazo class colour developing liquid 2.50ml, mixing, put 37 ℃ of water bath heat preservations 5 minutes, return to zero with the blank pipe, with 1cm optical path cuvette, mensuration is respectively managed absorbance A at 550--600nm wavelength place.
2. automatic analyzer analytical parameters reaction type: end-point method; Wavelength: 600nm; Temperature of reaction: 37 ℃; Pyridylazo class colour developing liquid: 250 μ l; Sample: 30 μ l; Reading duration: 300 seconds.
The content of serum levels of iron (μ mol/L)=17.90 * Au/As
Embodiment six
A kind of serum levels of iron rapid sensitive detection kit, this kit is made up of following reagent:
Colour developing liquid (pyridylazo class developer) 1 bottle
1 bottle of iron titer (concentration is 35.8 μ mol/L)
The preparation method of this kit realizes as follows:
(1) preparation method of pyridylazo class developer:
A, take by weighing 2-(5-bromo-2-pyridylazo)-5-(diethylamino phenol) 150mg,
B, the reagent among a is made into the stock solution of 100ml with absolute ethyl alcohol,
C, the stock solution 15ml that gets b add 15g Triton X-100 and 40g thiocarbamide, are diluted to 1000ml with the acetic acid sodium acetate aqueous solution of PH4.0.
(2) zinc titer: accurately take by weighing the metallic zinc (99.9%) of 100.0mg, the aqueous acetic acid dissolving back with an amount of 50% is diluted to 1000ml with distilled water, and dilute with water is the zinc titer of 50 times 35.80 μ mol/L again.
Detection method:
1. manual assay pipe (U), standard pipe (S), blank pipe (B) add serum, iron titer, each 0.3ml of distilled water respectively, every pipe adds pyridylazo class colour developing liquid 2.50ml, mixing, put 37 ℃ of water bath heat preservations 5 minutes, return to zero with the blank pipe, with 1cm optical path cuvette, mensuration is respectively managed absorbance A at 550--600nm wavelength place.
2. automatic analyzer analytical parameters reaction type: end-point method; Wavelength: 600nm; Temperature of reaction: 37 ℃; Pyridylazo class colour developing liquid: 250 μ l; Sample: 30 μ l; Reading duration: 300 seconds.
The content of serum levels of iron (μ mol/L)=35.80 * Au/As
Claims (9)
1, be used to detect the rapid sensitive detection kit of content of Cu in serum, it is characterized in that: it is made up of following reagent:
(1) oxygenant: concentration is hydrogen peroxide or the potassium persulfate of 30g/L;
(2) pyridylazo class colour developing liquid: solid content is by 20-100mg pyridylazo type organic in every liter of colour developing liquid, 5-20g surfactant and 20-100g screening agent are formed, liquid by pH=3.5-5.0 acetic acid-sodium acetate aqueous solution and absolute ethyl alcohol in 199: 1-19: 1 ratio is formed, described surfactant is that described surfactant is a polyoxyethylene laurel ether, polyoxyethylene iso-octyl phenyl ether, Triton X-100, polyoxyethylene sorbitan lauric acid monoester, polyoxyethylene sorbitan oleic acid monoester or polyvinylpyrrolidone, described screening agent are citric acid, sodium fluoride or sodium hexametaphosphate;
(3) copper titer: concentration is 15.74-31.48 μ mol/L.
2, the preparation method who is used to detect the rapid sensitive detection kit of content of Cu in serum according to claim 1, it is characterized in that: the preparation method of described pyridylazo class colour developing liquid is: every liter of colour developing liquid takes by weighing pyridylazo type organic 100-500mg, be made into the 100ml stock solution with absolute ethyl alcohol, get stock solution 5-20ml, the surfactant that adds 5-20g is diluted to 1000ml with acetic acid-sodium acetate solution of pH=3.5-5.0.
3, the rapid sensitive detection kit that is used to detect content of Cu in serum according to claim 1, it is characterized in that: described pyridylazo type organic is 2-(5-bromo-2-pyridylazo)-5-(diethylamino phenol), 2-(3,5-two bromo-2-pyridylazos)-and 5-diethylamino phenol, 2-(5-bromo-2-pyridylazo)-5-dimethylamino phenol, 2-(3,5-two bromo-2-pyridylazos)-5-dimethylamino phenol or 2-(5-nitro-2-pyridylazo)-5-[N-n-pro-pyl-N-(3-sulfonic acid propyl group) amino] the phenol disodium.
4, be used to detect the rapid sensitive detection kit of Zn in serum content, it is characterized in that: it is made up of following reagent:
(1) pyridylazo class colour developing liquid: solid content is by 20-100mg pyridylazo type organic in every liter of colour developing liquid, 5-20g surfactant and 20-100g screening agent are formed, liquid by the Tri(Hydroxymethyl) Amino Methane Hydrochloride aqueous solution of pH=8.0-10 and absolute ethyl alcohol in 199: 1-19: 1 ratio is formed, described surfactant is a polyoxyethylene iso-octyl phenyl ether, Triton X-100, polyoxyethylene sorbitan lauric acid monoester, polyoxyethylene sorbitan oleic acid monoester or polyvinylpyrrolidone, described screening agent are cupferron, citric acid, mercaptoacetic acid, thiocarbamide or sodium hexametaphosphate;
(2) zinc titer: concentration is 15.3-30.6 μ mol/L.
5, the preparation method who is used to detect the rapid sensitive detection kit of Zn in serum content according to claim 4, it is characterized in that: the preparation method of described pyridylazo class colour developing liquid is: every liter of colour developing liquid takes by weighing the screening agent of pyridylazo type organic 100-500mg and 20-100g, be made into the 100ml stock solution with absolute ethyl alcohol, get stock solution 5-50ml, the surfactant that adds 5-20g is diluted to 1000ml with the Tri(Hydroxymethyl) Amino Methane Hydrochloride aqueous solution of pH=8.0-10.0.
6, the rapid sensitive detection kit that is used to detect Zn in serum content according to claim 4: described pyridylazo type organic is 2-(5-bromo-2-pyridylazo)-5-(diethylamino phenol), 2-(3,5-two bromo-2-pyridylazos)-5-diethylamino phenol, 2-(5-bromo-2-pyridylazo)-5-dimethylamino phenol, 2-(3,5-two bromo-2-pyridylazos)-5-dimethylamino phenol (be called for short 3,5-diBr-DMPAP) or 2-(5-nitro-2-pyridylazo)-5-[N-n-pro-pyl-N-(3-sulfonic acid propyl group) amino] the phenol disodium.
7, be used for detecting the rapid sensitive detection kit of serum iron content, it is characterized in that: it is made up of following reagent:
(1) pyridylazo class colour developing liquid: solid content is made up of 20-100mg pyridylazo type organic, 5-20g surfactant and 20-100g screening agent in every liter of colour developing liquid, liquid by pH=3.5-5.0 acetic acid-sodium acetate aqueous solution and absolute ethyl alcohol in 199: 1-19: 1 ratio is formed, described surfactant is polyoxyethylene iso-octyl phenyl ether, Triton X-100, polyoxyethylene sorbitan lauric acid monoester, polyoxyethylene sorbitan oleic acid monoester or polyvinylpyrrolidone, and described screening agent is mercaptoacetic acid or thiocarbamide;
(2) iron titer: concentration is 17.9-35.8 μ mol/L.
8, the preparation method who is used for detecting the rapid sensitive detection kit of serum iron content according to claim 7, it is characterized in that: the preparation method of described pyridylazo class colour developing liquid is: take by weighing pyridylazo type organic 100-500mg and 20-100g screening agent, be made into the 100ml stock solution with absolute ethyl alcohol, get stock solution 5-50ml, the surfactant that adds 5-20g is diluted to 1000ml with acetic acid-sodium acetate solution of pH=3.5-5.0.
9, the rapid sensitive detection kit that is used for detecting the serum iron content according to claim 7, it is characterized in that: described pyridylazo type organic is 2-(5-bromo-2-pyridylazo)-5-(diethylamino phenol), 2-(3,5-two bromo-2-pyridylazos)-and 5-diethylamino phenol, 2-(5-bromo-2-pyridylazo)-5-dimethylamino phenol, 2-(3,5-two bromo-2-pyridylazos)-5-dimethylamino phenol or 2-(5-nitro-2-pyridylazo)-5-[N-n-pro-pyl-N-(3-sulfonic acid propyl group) amino] the phenol disodium.
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| CN100501383C (en) * | 2006-09-20 | 2009-06-17 | 浙江大学 | Method for fast determination of iron content in rice seed |
| CN102053068A (en) * | 2010-11-02 | 2011-05-11 | 首都医科大学附属北京朝阳医院 | Serum iron content detection method and special standard substance thereof |
| CN102175676A (en) * | 2011-01-20 | 2011-09-07 | 东华理工大学 | Novel developing body for measuring copper ions by spectrophotometry and preparation process thereof |
| CN102353671A (en) * | 2011-06-27 | 2012-02-15 | 董理 | Method and kit for detecting serium inorganic phosphorus |
| CN103364354A (en) * | 2013-06-26 | 2013-10-23 | 斯涛利科技发展(天津)有限公司 | Method for colorimetric detection of hydrogen peroxide by utilizing metal organic complexes |
| CN103728228A (en) * | 2012-10-10 | 2014-04-16 | 三星Sdi株式会社 | Method for detecting non-magnetic metal particles contained in secondary battery materials |
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| CN105181692A (en) * | 2015-09-30 | 2015-12-23 | 山东博科生物产业有限公司 | Colorimetry detection kit for IMA (ischemia modified albumin) |
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| CN109883972A (en) * | 2019-04-11 | 2019-06-14 | 哈尔滨工业大学（威海） | Dissociate Methods For The Determination of Iron in a kind of red blood cell |
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| CN102053068A (en) * | 2010-11-02 | 2011-05-11 | 首都医科大学附属北京朝阳医院 | Serum iron content detection method and special standard substance thereof |
| CN102053068B (en) * | 2010-11-02 | 2012-10-24 | 首都医科大学附属北京朝阳医院 | Serum iron content detection method and special standard substance thereof |
| CN102175676A (en) * | 2011-01-20 | 2011-09-07 | 东华理工大学 | Novel developing body for measuring copper ions by spectrophotometry and preparation process thereof |
| CN102353671A (en) * | 2011-06-27 | 2012-02-15 | 董理 | Method and kit for detecting serium inorganic phosphorus |
| CN103728228A (en) * | 2012-10-10 | 2014-04-16 | 三星Sdi株式会社 | Method for detecting non-magnetic metal particles contained in secondary battery materials |
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