CN111157431A - Reagent for classifying nucleated erythrocytes and basophils - Google Patents
Reagent for classifying nucleated erythrocytes and basophils Download PDFInfo
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- CN111157431A CN111157431A CN202010035805.5A CN202010035805A CN111157431A CN 111157431 A CN111157431 A CN 111157431A CN 202010035805 A CN202010035805 A CN 202010035805A CN 111157431 A CN111157431 A CN 111157431A
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- blood cells
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- 239000003153 chemical reaction reagent Substances 0.000 title claims abstract description 69
- 210000003651 basophil Anatomy 0.000 title claims abstract description 37
- 210000003924 normoblast Anatomy 0.000 title abstract description 15
- 239000004094 surface-active agent Substances 0.000 claims abstract description 29
- 210000003743 erythrocyte Anatomy 0.000 claims abstract description 28
- 150000001413 amino acids Chemical class 0.000 claims abstract description 23
- 239000000022 bacteriostatic agent Substances 0.000 claims abstract description 10
- 239000003093 cationic surfactant Substances 0.000 claims abstract description 10
- 239000000049 pigment Substances 0.000 claims abstract description 7
- 150000001298 alcohols Chemical class 0.000 claims abstract description 3
- 239000000872 buffer Substances 0.000 claims abstract 2
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 claims description 18
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 13
- -1 alkyl betaine Chemical compound 0.000 claims description 13
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 12
- 229920003171 Poly (ethylene oxide) Polymers 0.000 claims description 11
- 239000007850 fluorescent dye Substances 0.000 claims description 8
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 6
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 6
- MUBZPKHOEPUJKR-UHFFFAOYSA-N Oxalic acid Chemical compound OC(=O)C(O)=O MUBZPKHOEPUJKR-UHFFFAOYSA-N 0.000 claims description 6
- 239000007853 buffer solution Substances 0.000 claims description 6
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims description 6
- 150000001875 compounds Chemical class 0.000 claims description 6
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 4
- 150000005215 alkyl ethers Chemical class 0.000 claims description 4
- WPYMKLBDIGXBTP-UHFFFAOYSA-N benzoic acid Chemical compound OC(=O)C1=CC=CC=C1 WPYMKLBDIGXBTP-UHFFFAOYSA-N 0.000 claims description 4
- 235000014113 dietary fatty acids Nutrition 0.000 claims description 4
- 229930195729 fatty acid Natural products 0.000 claims description 4
- 239000000194 fatty acid Substances 0.000 claims description 4
- 150000004665 fatty acids Chemical class 0.000 claims description 4
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 claims description 4
- XNGIFLGASWRNHJ-UHFFFAOYSA-N phthalic acid Chemical compound OC(=O)C1=CC=CC=C1C(O)=O XNGIFLGASWRNHJ-UHFFFAOYSA-N 0.000 claims description 4
- YGSDEFSMJLZEOE-UHFFFAOYSA-N salicylic acid Chemical compound OC(=O)C1=CC=CC=C1O YGSDEFSMJLZEOE-UHFFFAOYSA-N 0.000 claims description 4
- IWIUXJGIDSGWDN-UQKRIMTDSA-M sodium;(2s)-2-(dodecanoylamino)pentanedioate;hydron Chemical group [Na+].CCCCCCCCCCCC(=O)N[C@H](C([O-])=O)CCC(O)=O IWIUXJGIDSGWDN-UQKRIMTDSA-M 0.000 claims description 4
- 150000001408 amides Chemical class 0.000 claims description 3
- 229940051841 polyoxyethylene ether Drugs 0.000 claims description 3
- 229920000056 polyoxyethylene ether Polymers 0.000 claims description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 3
- TUBPSFQENHCYBW-HVDRVSQOSA-N (2s)-2-aminopentanedioic acid;2-[bis(2-hydroxyethyl)amino]ethanol Chemical compound OC(=O)[C@@H](N)CCC(O)=O.OCCN(CCO)CCO TUBPSFQENHCYBW-HVDRVSQOSA-N 0.000 claims description 2
- BJEPYKJPYRNKOW-REOHCLBHSA-N (S)-malic acid Chemical compound OC(=O)[C@@H](O)CC(O)=O BJEPYKJPYRNKOW-REOHCLBHSA-N 0.000 claims description 2
- ZIRURAJAJIQZFG-UHFFFAOYSA-N 1-aminopropane-1-sulfonic acid Chemical compound CCC(N)S(O)(=O)=O ZIRURAJAJIQZFG-UHFFFAOYSA-N 0.000 claims description 2
- QCDWFXQBSFUVSP-UHFFFAOYSA-N 2-phenoxyethanol Chemical group OCCOC1=CC=CC=C1 QCDWFXQBSFUVSP-UHFFFAOYSA-N 0.000 claims description 2
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 claims description 2
- 239000005711 Benzoic acid Substances 0.000 claims description 2
- KWIUHFFTVRNATP-UHFFFAOYSA-N Betaine Natural products C[N+](C)(C)CC([O-])=O KWIUHFFTVRNATP-UHFFFAOYSA-N 0.000 claims description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical group [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 2
- FEWJPZIEWOKRBE-JCYAYHJZSA-N Dextrotartaric acid Chemical compound OC(=O)[C@H](O)[C@@H](O)C(O)=O FEWJPZIEWOKRBE-JCYAYHJZSA-N 0.000 claims description 2
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 claims description 2
- OFOBLEOULBTSOW-UHFFFAOYSA-N Propanedioic acid Natural products OC(=O)CC(O)=O OFOBLEOULBTSOW-UHFFFAOYSA-N 0.000 claims description 2
- ABBQHOQBGMUPJH-UHFFFAOYSA-M Sodium salicylate Chemical compound [Na+].OC1=CC=CC=C1C([O-])=O ABBQHOQBGMUPJH-UHFFFAOYSA-M 0.000 claims description 2
- KDYFGRWQOYBRFD-UHFFFAOYSA-N Succinic acid Natural products OC(=O)CCC(O)=O KDYFGRWQOYBRFD-UHFFFAOYSA-N 0.000 claims description 2
- FEWJPZIEWOKRBE-UHFFFAOYSA-N Tartaric acid Natural products [H+].[H+].[O-]C(=O)C(O)C(O)C([O-])=O FEWJPZIEWOKRBE-UHFFFAOYSA-N 0.000 claims description 2
- 150000003973 alkyl amines Chemical class 0.000 claims description 2
- 125000000217 alkyl group Chemical group 0.000 claims description 2
- BJEPYKJPYRNKOW-UHFFFAOYSA-N alpha-hydroxysuccinic acid Natural products OC(=O)C(O)CC(O)=O BJEPYKJPYRNKOW-UHFFFAOYSA-N 0.000 claims description 2
- 150000001450 anions Chemical class 0.000 claims description 2
- 235000010233 benzoic acid Nutrition 0.000 claims description 2
- 229960003237 betaine Drugs 0.000 claims description 2
- KDYFGRWQOYBRFD-NUQCWPJISA-N butanedioic acid Chemical compound O[14C](=O)CC[14C](O)=O KDYFGRWQOYBRFD-NUQCWPJISA-N 0.000 claims description 2
- 125000004432 carbon atom Chemical group C* 0.000 claims description 2
- 239000004359 castor oil Substances 0.000 claims description 2
- 235000019438 castor oil Nutrition 0.000 claims description 2
- 229910001914 chlorine tetroxide Inorganic materials 0.000 claims description 2
- DDXLVDQZPFLQMZ-UHFFFAOYSA-M dodecyl(trimethyl)azanium;chloride Chemical group [Cl-].CCCCCCCCCCCC[N+](C)(C)C DDXLVDQZPFLQMZ-UHFFFAOYSA-M 0.000 claims description 2
- 150000002148 esters Chemical class 0.000 claims description 2
- 150000002191 fatty alcohols Chemical class 0.000 claims description 2
- 235000019253 formic acid Nutrition 0.000 claims description 2
- ZEMPKEQAKRGZGQ-XOQCFJPHSA-N glycerol triricinoleate Natural products CCCCCC[C@@H](O)CC=CCCCCCCCC(=O)OC[C@@H](COC(=O)CCCCCCCC=CC[C@@H](O)CCCCCC)OC(=O)CCCCCCCC=CC[C@H](O)CCCCCC ZEMPKEQAKRGZGQ-XOQCFJPHSA-N 0.000 claims description 2
- MGIYRDNGCNKGJU-UHFFFAOYSA-N isothiazolinone Chemical compound O=C1C=CSN1 MGIYRDNGCNKGJU-UHFFFAOYSA-N 0.000 claims description 2
- VZCYOOQTPOCHFL-UPHRSURJSA-N maleic acid Chemical compound OC(=O)\C=C/C(O)=O VZCYOOQTPOCHFL-UPHRSURJSA-N 0.000 claims description 2
- 239000011976 maleic acid Substances 0.000 claims description 2
- 239000001630 malic acid Substances 0.000 claims description 2
- 235000011090 malic acid Nutrition 0.000 claims description 2
- LPUQAYUQRXPFSQ-DFWYDOINSA-M monosodium L-glutamate Chemical compound [Na+].[O-]C(=O)[C@@H](N)CCC(O)=O LPUQAYUQRXPFSQ-DFWYDOINSA-M 0.000 claims description 2
- 235000006408 oxalic acid Nutrition 0.000 claims description 2
- FJKROLUGYXJWQN-UHFFFAOYSA-N papa-hydroxy-benzoic acid Natural products OC(=O)C1=CC=C(O)C=C1 FJKROLUGYXJWQN-UHFFFAOYSA-N 0.000 claims description 2
- VLTRZXGMWDSKGL-UHFFFAOYSA-M perchlorate Chemical compound [O-]Cl(=O)(=O)=O VLTRZXGMWDSKGL-UHFFFAOYSA-M 0.000 claims description 2
- 229960005323 phenoxyethanol Drugs 0.000 claims description 2
- 229920002503 polyoxyethylene-polyoxypropylene Polymers 0.000 claims description 2
- IWZKICVEHNUQTL-UHFFFAOYSA-M potassium hydrogen phthalate Chemical compound [K+].OC(=O)C1=CC=CC=C1C([O-])=O IWZKICVEHNUQTL-UHFFFAOYSA-M 0.000 claims description 2
- 150000003242 quaternary ammonium salts Chemical group 0.000 claims description 2
- 229960004889 salicylic acid Drugs 0.000 claims description 2
- 229940071089 sarcosinate Drugs 0.000 claims description 2
- 239000011734 sodium Substances 0.000 claims description 2
- 229910052708 sodium Inorganic materials 0.000 claims description 2
- 239000001509 sodium citrate Substances 0.000 claims description 2
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 claims description 2
- 229940079781 sodium cocoyl glutamate Drugs 0.000 claims description 2
- 229940045944 sodium lauroyl glutamate Drugs 0.000 claims description 2
- 229960004025 sodium salicylate Drugs 0.000 claims description 2
- 229940074404 sodium succinate Drugs 0.000 claims description 2
- ZDQYSKICYIVCPN-UHFFFAOYSA-L sodium succinate (anhydrous) Chemical compound [Na+].[Na+].[O-]C(=O)CCC([O-])=O ZDQYSKICYIVCPN-UHFFFAOYSA-L 0.000 claims description 2
- LFUXMTKAILZVTA-ZOWNYOTGSA-M sodium;(2s)-2-(dodecanoylamino)propanoate Chemical compound [Na+].CCCCCCCCCCCC(=O)N[C@@H](C)C([O-])=O LFUXMTKAILZVTA-ZOWNYOTGSA-M 0.000 claims description 2
- XNRNJIIJLOFJEK-UHFFFAOYSA-N sodium;1-oxidopyridine-2-thione Chemical compound [Na+].[O-]N1C=CC=CC1=S XNRNJIIJLOFJEK-UHFFFAOYSA-N 0.000 claims description 2
- 239000011975 tartaric acid Substances 0.000 claims description 2
- 235000002906 tartaric acid Nutrition 0.000 claims description 2
- VZCYOOQTPOCHFL-UHFFFAOYSA-N trans-butenedioic acid Natural products OC(=O)C=CC(O)=O VZCYOOQTPOCHFL-UHFFFAOYSA-N 0.000 claims description 2
- 229940048912 triethanolamine cocoyl glutamate Drugs 0.000 claims description 2
- CEYYIKYYFSTQRU-UHFFFAOYSA-M trimethyl(tetradecyl)azanium;chloride Chemical compound [Cl-].CCCCCCCCCCCCCC[N+](C)(C)C CEYYIKYYFSTQRU-UHFFFAOYSA-M 0.000 claims description 2
- 235000001014 amino acid Nutrition 0.000 claims 3
- 239000003795 chemical substances by application Substances 0.000 claims 3
- 210000004369 blood Anatomy 0.000 abstract description 15
- 239000008280 blood Substances 0.000 abstract description 15
- 210000000265 leukocyte Anatomy 0.000 abstract description 9
- 239000000126 substance Substances 0.000 abstract description 8
- 230000000844 anti-bacterial effect Effects 0.000 abstract description 5
- 210000003714 granulocyte Anatomy 0.000 abstract description 5
- 238000000338 in vitro Methods 0.000 abstract description 4
- 239000003242 anti bacterial agent Substances 0.000 abstract description 2
- 238000004043 dyeing Methods 0.000 abstract description 2
- 238000001556 precipitation Methods 0.000 abstract description 2
- 239000000243 solution Substances 0.000 description 5
- 210000004027 cell Anatomy 0.000 description 3
- 201000010099 disease Diseases 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 description 2
- 230000007794 irritation Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 208000032791 BCR-ABL1 positive chronic myelogenous leukemia Diseases 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- 208000010833 Chronic myeloid leukaemia Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
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- 208000017604 Hodgkin disease Diseases 0.000 description 1
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- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 241000700159 Rattus Species 0.000 description 1
- 239000012445 acidic reagent Substances 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000004820 blood count Methods 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
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- 231100000135 cytotoxicity Toxicity 0.000 description 1
- 238000005202 decontamination Methods 0.000 description 1
- 230000003588 decontaminative effect Effects 0.000 description 1
- 239000007857 degradation product Substances 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
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- 238000010586 diagram Methods 0.000 description 1
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- 231100000206 health hazard Toxicity 0.000 description 1
- 208000010501 heavy metal poisoning Diseases 0.000 description 1
- 239000003219 hemolytic agent Substances 0.000 description 1
- 210000004400 mucous membrane Anatomy 0.000 description 1
- 231100000017 mucous membrane irritation Toxicity 0.000 description 1
- 231100000252 nontoxic Toxicity 0.000 description 1
- 230000003000 nontoxic effect Effects 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 210000005259 peripheral blood Anatomy 0.000 description 1
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Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
- G01N15/10—Investigating individual particles
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N2015/1006—Investigating individual particles for cytology
-
- G01N2015/1022—
-
- G01N2015/1028—
Abstract
The invention relates to a reagent for classifying nucleated red blood cells and basophilic granulocyte, belonging to the technical field of in vitro diagnostic reagents. The reagent for classifying nucleated erythrocytes and basophils comprises: a first reagent and a second reagent; the first reagent comprises: buffer, amino acid surfactant, cationic surfactant, and bacteriostatic agent; the second reagent comprises: fluorescent pigments, and alcohols. The invention adopts the amino acid surfactant to replace the traditional surfactant, provides the reagent which can accurately distinguish and count the nucleated erythrocytes and basophilic granulocytes in the whole blood sample from other white blood cells, not only increases the safety of producers and users, but also improves the solubility of pigment substances, reduces the precipitation of the pigment substances and improves the stability of the dyeing reagent due to the excellent emulsibility of the amino acid surfactant. In addition, the amino acid surfactant has a certain antibacterial effect, and can reduce the use of antibacterial agents or bacteriostatic agents.
Description
Technical Field
The invention belongs to the technical field of in-vitro diagnostic reagents, and particularly relates to a reagent for classifying nucleated erythrocytes and basophilic granulocytes.
Background
Human normal mature red blood cells do not contain a nucleus. However, in special cases, including neonates or certain pathological conditions, human peripheral blood can exhibit nucleated red blood cells, a type of naive red blood cells. The detection of nucleated red blood cells can provide a basis for the diagnosis of certain diseases.
In normal human body, basophils are small in number, and only account for 0-1% of the total number of leukocytes. However, in diseases such as heavy metal poisoning, Hodgkin's disease, chronic myelocytic leukemia and the like, the number of basophils is obviously increased, so that the basophil count has important significance for diagnosing the diseases.
Both basophils and nucleated erythrocytes can be measured by treating a blood sample with an acidic reagent, mixing the sample with an acidic hemolytic agent, removing normal erythrocytes, adding a fluorescent staining solution for staining, and counting the number and percentage of nucleated erythrocytes and basophils by measuring the fluorescence intensity and the scattered light intensity.
The traditional surfactants such as sodium dodecyl sulfate and the like have strong irritation and cytotoxicity to eyes, respiratory tracts and mucous membranes of people, and have certain health hazard to producers and users of in vitro diagnostic reagents. Meanwhile, the traditional surfactant seriously pollutes water sources and is not environment-friendly.
The research of amino acid type surfactants starts more than 40 years ago, and the biomass-based surfactants have high safety to the environment and organisms due to the amino acid skeleton structure in molecules, and have some unique functions besides the common characteristics of the surfactants. 1. The surface activity is excellent; the salt surfactant has excellent functions of decontamination, calcium resistance, foaming and emulsification. 2. The biodegradability is good; the amino acid surfactant takes renewable substances as raw materials, has good biodegradability, and can be decomposed into fatty acid and amino acid by enzyme in human body. 3. The safety is high; researchers have performed subacute tests, chronic toxicity tests, mucosal irritation tests and the like on white rats and rabbits, and the results show that the N-acyl amino acid has smaller irritation and higher safety than sodium dodecyl sulfate. 4. The antibacterial ability is strong; the amino acid type surfactant has antibacterial effect on gram-positive bacteria and partial filamentous bacteria. In conclusion, the amino acid type surfactant has the characteristics of good wettability, foamability, antibacterial property, corrosion resistance, antistatic property and the like, is nontoxic and harmless, is mild to skin, has no influence on environment because degradation products are amino acid and fatty acid, and has good compatibility with other surfactants. At present, the amino acid surfactant is mainly widely used in daily chemical products, and no relevant report is available for in vitro diagnostic reagents.
Disclosure of Invention
The invention aims to solve the technical problems in the prior art, and provides a reagent for classifying nucleated erythrocytes and basophils by replacing the traditional surfactant with an amino acid surfactant. The reagent provided by the invention can accurately distinguish and count nucleated erythrocytes and basophils in a whole blood sample from other leukocytes.
In order to solve the technical problems, the technical scheme of the invention is as follows:
the invention provides a reagent for classifying nucleated erythrocytes and basophils, which comprises the following components: a first reagent and a second reagent;
the first reagent comprises: a buffer solution with a pH value of 2-4, an amino acid surfactant, a cationic surfactant and a bacteriostatic agent;
the second reagent comprises: fluorescent pigments, and alcohols.
In the above technical scheme, the buffer solution is selected from one or more of citric acid, sodium citrate, benzoic acid, potassium hydrogen phthalate, phthalic acid, salicylic acid, sodium salicylate, malic acid, tartaric acid, oxalic acid, succinic acid, sodium succinate, maleic acid, formic acid, and acetic acid.
In the above technical scheme, the amino acid surfactant is sodium lauroyl glutamate, sodium cocoyl glutamate, TEA cocoyl glutamate, sodium N-lauroyl alaninate, sodium N-lauroyl glutamate, sodium N-fatty acyl sarcosinate, alkyl betaine, aminopropanesulfonate, polyoxyethylene hydrogenated castor oil, polyoxyethylene alkyl ether, polyoxyethylene polyoxypropylene alkyl ether, polyoxyethylene alkylamine, alkylphenol polyoxyethylene, fatty alcohol polyoxyethylene ether, fatty acid polyoxyethylene ester, alkylol amide, or polyoxyethylene amide.
In the above technical scheme, the cationic surfactant is selected from quaternary ammonium salts with carbon chains of 8-14.
In the above technical solution, the cationic surfactant is selected from dodecyl trimethyl ammonium chloride or tetradecyl trimethyl ammonium chloride.
In the technical scheme, the bacteriostatic agent is selected from phenoxyethanol, isothiazolinone or sodium pyrithione.
In the above technical solution, the structural formula of the fluorescent dye is shown as general formula (I):
in the formula, X-Is an anion selected from F-、Cl-、Br-、I-、CF3SO3 -、BF4 -Or ClO4 -；
R1、R2Are the same or different alkyl groups;
R3、R4the following structures, which may be the same or different:
in the above technical solution, the fluorescent dye is selected from compounds represented by any one of the following structures:
in the above technical scheme, the alcohol is an alcohol with 1-4 carbon atoms, and is selected from one or more of methanol, ethanol, ethylene glycol and glycerol.
In the above technical solution, the reagent for classifying nucleated red blood cells and basophils comprises the following components:
1L of water;
the first reagent comprises:
buffer solution: 1.5-2.9 g;
amino acid-based surfactant: 300-500 mg;
cationic surfactant: 0.5-1.6 g;
bacteriostatic agent: 300-;
the second reagent comprises:
fluorescent dye: 5 mg;
alcohol: 100 mL;
the pH of the reagent was adjusted to 2.8-3.0 using 1M hydrochloric acid.
The invention has the beneficial effects that:
the invention adopts the amino acid surfactant to replace the traditional surfactant, provides the reagent which can accurately distinguish and count the nucleated erythrocytes and basophilic granulocytes in the whole blood sample from other white blood cells, not only increases the safety of producers and users, but also improves the solubility of pigment substances, reduces the precipitation of the pigment substances and improves the stability of the dyeing reagent due to the excellent emulsibility of the amino acid surfactant. In addition, the amino acid surfactant has a certain antibacterial effect, and can reduce the use of antibacterial agents or bacteriostatic agents.
Drawings
The present invention will be described in further detail with reference to the accompanying drawings and specific embodiments.
FIG. 1 is a graph showing the results of the reagent of example 1 of the present invention used for classifying nucleated red blood cells and basophils in a whole blood sample.
FIG. 2 is a graph showing the results of the reagent of example 2 of the present invention used for classifying nucleated red blood cells and basophils in a whole blood sample.
FIG. 3 is a graph showing the results of the reagent of example 3 of the present invention used for classifying nucleated red blood cells and basophils in a whole blood sample.
FIG. 4 is a graph showing the results of the reagent of example 4 of the present invention used for classifying nucleated red blood cells and basophils in a whole blood sample.
Detailed Description
Example 1
The reagent is prepared according to the following contents of components:
a first reagent:
a second reagent:
compound of Structure (II) 5mg
Ethylene glycol 100mL
Example 2
The reagent is prepared according to the following contents of components:
a first reagent:
a second reagent:
compound of Structure (III) 8mg
Ethylene glycol 100mL
Example 3
The reagent is prepared according to the following contents of components:
a first reagent:
a second reagent:
compound of Structure (IV) 5mg
Methanol 3mL
Ethylene glycol 97mL
Example 4
The reagent is prepared according to the following contents of components:
a first reagent:
a second reagent:
compound of structure (V) 5mg
Methanol 3mL
Ethylene glycol 97mL
In the above embodiments, the buffer solution, the amino acid surfactant, the cationic surfactant, and the bacteriostatic agent in the first reagent may be replaced by any one of the substances within the above-mentioned limits, and the fluorescent dye and the alcohol in the second reagent may be replaced by any one of the substances within the above-mentioned limits, which are not exemplified herein.
Example 5
The reagents prepared in the embodiments 1 to 4 of the present invention are respectively used for classifying nucleated erythrocytes and basophilic granulocytes in whole blood samples, and the specific steps are as follows:
after 1mL of the first reagent, 20. mu.L of the second reagent, and 18. mu.L of the whole blood sample were thoroughly mixed and reacted at 41 ℃ for 12 seconds, fluorescence and forward scattered light signals of leukocytes and nucleated erythrocytes in the sample were measured using an automatic blood cell counting apparatus. And drawing a scatter diagram with the forward scattered light intensity and the fluorescence intensity as two axes.
FIG. 1 is a graph showing the results of the reagent of example 1 of the present invention used for classifying nucleated red blood cells and basophils in a whole blood sample. From this figure, it can be seen that: treatment of a whole blood sample with the reagent of example 1 clearly distinguishes basophils from leukocytes other than basophils.
FIG. 2 is a graph showing the results of the reagent of example 2 of the present invention used for classifying nucleated red blood cells and basophils in a whole blood sample. From this figure, it can be seen that: when cells appearing in a predetermined area in the scattergram are basophils, the number of basophils and the ratio of basophils to the total number of leukocytes can be accurately determined.
FIG. 3 is a graph showing the results of the reagent of example 3 of the present invention used for classifying nucleated red blood cells and basophils in a whole blood sample. From this figure, it can be seen that: when the cells appearing in a predetermined region in the scattergram are nucleated red blood cells, the number of nucleated red blood cells and the ratio of the number of nucleated red blood cells to the total number of white blood cells can be accurately determined.
FIG. 4 is a graph showing the results of the reagent of example 4 of the present invention used for classifying nucleated red blood cells and basophils in a whole blood sample. From this figure, it can be seen that: the number of basophils and nucleated erythrocytes and the ratio thereof to the total number of leukocytes can be accurately determined by using cells appearing in a predetermined region in the scattergram as basophils and nucleated erythrocytes.
It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications therefrom are within the scope of the invention.
Claims (10)
1. An agent for classifying nucleated red blood cells and basophils, comprising: a first reagent and a second reagent;
the first reagent comprises: a buffer solution with a pH value of 2-4, an amino acid surfactant, a cationic surfactant and a bacteriostatic agent;
the second reagent comprises: fluorescent pigments, and alcohols.
2. The reagent for nucleated red blood cells, basophils classification of claim 1, wherein said buffer is selected from one or more of citric acid, sodium citrate, benzoic acid, potassium hydrogen phthalate, phthalic acid, salicylic acid, sodium salicylate, malic acid, tartaric acid, oxalic acid, succinic acid, sodium succinate, maleic acid, formic acid, and acetic acid.
3. An agent for nucleated red blood cells, basophils according to claim 1, wherein said amino acid-based surfactant is sodium lauroyl glutamate, sodium cocoyl glutamate, TEA cocoyl glutamate, sodium N-lauroyl alanine, sodium N-lauroyl glutamate, sodium N-fatty acyl sarcosinate, alkyl betaine, aminopropanesulfonate, polyoxyethylene hydrogenated castor oil, polyoxyethylene alkyl ether, polyoxyethylene polyoxypropylene alkyl ether, polyoxyethylene alkylamine, alkylphenol polyoxyethylene ether, fatty alcohol polyoxyethylene ether, fatty acid polyoxyethylene ester, alkylolamide, or polyoxyethylene amide.
4. The reagent for the classification of nucleated red blood cells and basophils according to claim 1, wherein said cationic surfactant is selected from quaternary ammonium salts having a carbon chain of 8-14.
5. The reagent for the classification of nucleated red blood cells, basophils according to claim 4, wherein said cationic surfactant is selected from dodecyltrimethylammonium chloride, or tetradecyltrimethylammonium chloride.
6. The agent for nucleated red blood cell, basophil classification according to claim 1, wherein said bacteriostatic agent is selected from phenoxyethanol, isothiazolinone, or sodium pyrithione.
7. The reagent for classifying nucleated red blood cells and basophils according to claim 1, wherein the structural formula of the fluorescent dye is represented by general formula (I):
in the formula, X-Is an anion selected from F-、Cl-、Br-、I-、CF3SO3 -、BF4 -Or ClO4 -；
R1、R2Are the same or different alkyl groups;
R3、R4the following structures, which may be the same or different:
8. the reagent for classifying nucleated red blood cells and basophils according to claim 7, wherein the fluorescent dye is selected from compounds represented by any one of the following structures:
9. the reagent for classifying nucleated red blood cells and basophils according to claim 1, wherein the alcohol is an alcohol having 1 to 4 carbon atoms and is one or more selected from the group consisting of methanol, ethanol, ethylene glycol and glycerol.
10. The reagent for classifying nucleated red blood cells and basophils according to any one of claims 1 to 9, wherein the reagent comprises the following components in percentage by weight:
1L of water;
the first reagent comprises:
buffer solution: 1.5-2.9 g;
amino acid-based surfactant: 300-500 mg;
cationic surfactant: 0.5-1.6 g;
bacteriostatic agent: 300-;
the second reagent comprises:
fluorescent dye: 5 mg;
alcohol: 100 mL;
the pH of the reagent was adjusted to 2.8-3.0 using 1M hydrochloric acid.
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Application publication date: 20200515 |