CN108148887A - Coloring composition for detecting microorganism infection and preparation method thereof - Google Patents
Coloring composition for detecting microorganism infection and preparation method thereof Download PDFInfo
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- CN108148887A CN108148887A CN201810048667.7A CN201810048667A CN108148887A CN 108148887 A CN108148887 A CN 108148887A CN 201810048667 A CN201810048667 A CN 201810048667A CN 108148887 A CN108148887 A CN 108148887A
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- coloring composition
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- microorganism
- sample
- tissue
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- 239000000203 mixture Substances 0.000 title claims abstract description 99
- 238000004040 coloring Methods 0.000 title claims abstract description 93
- 244000005700 microbiome Species 0.000 title claims abstract description 50
- 208000015181 infectious disease Diseases 0.000 title claims abstract description 31
- 238000002360 preparation method Methods 0.000 title abstract description 18
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims abstract description 44
- GEHJYWRUCIMESM-UHFFFAOYSA-L sodium sulfite Chemical compound [Na+].[Na+].[O-]S([O-])=O GEHJYWRUCIMESM-UHFFFAOYSA-L 0.000 claims abstract description 42
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims abstract description 40
- 238000001514 detection method Methods 0.000 claims abstract description 30
- 229960000907 methylthioninium chloride Drugs 0.000 claims abstract description 27
- 239000012530 fluid Substances 0.000 claims abstract description 25
- LSNNMFCWUKXFEE-UHFFFAOYSA-N Sulfurous acid Chemical class OS(O)=O LSNNMFCWUKXFEE-UHFFFAOYSA-N 0.000 claims abstract description 23
- 235000010265 sodium sulphite Nutrition 0.000 claims abstract description 21
- 239000003638 chemical reducing agent Substances 0.000 claims abstract description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 19
- 150000001720 carbohydrates Chemical class 0.000 claims abstract description 17
- LSNNMFCWUKXFEE-UHFFFAOYSA-L sulfite Chemical compound [O-]S([O-])=O LSNNMFCWUKXFEE-UHFFFAOYSA-L 0.000 claims abstract description 17
- 230000002757 inflammatory effect Effects 0.000 claims abstract description 14
- 230000003902 lesion Effects 0.000 claims abstract description 14
- RBTBFTRPCNLSDE-UHFFFAOYSA-N 3,7-bis(dimethylamino)phenothiazin-5-ium Chemical compound C1=CC(N(C)C)=CC2=[S+]C3=CC(N(C)C)=CC=C3N=C21 RBTBFTRPCNLSDE-UHFFFAOYSA-N 0.000 claims abstract 4
- 238000000034 method Methods 0.000 claims description 35
- 239000000243 solution Substances 0.000 claims description 25
- 241001502500 Trichomonadida Species 0.000 claims description 24
- 210000001519 tissue Anatomy 0.000 claims description 23
- 235000014633 carbohydrates Nutrition 0.000 claims description 18
- 238000004043 dyeing Methods 0.000 claims description 16
- 241000222122 Candida albicans Species 0.000 claims description 15
- 230000008859 change Effects 0.000 claims description 15
- 241000193830 Bacillus <bacterium> Species 0.000 claims description 13
- 239000012153 distilled water Substances 0.000 claims description 13
- 241000894006 Bacteria Species 0.000 claims description 12
- 241000283707 Capra Species 0.000 claims description 12
- 241000282693 Cercopithecidae Species 0.000 claims description 12
- 241000283073 Equus caballus Species 0.000 claims description 12
- 241000282898 Sus scrofa Species 0.000 claims description 12
- 241000124008 Mammalia Species 0.000 claims description 11
- 206010007134 Candida infections Diseases 0.000 claims description 10
- 229920000742 Cotton Polymers 0.000 claims description 10
- 238000001574 biopsy Methods 0.000 claims description 10
- 201000003984 candidiasis Diseases 0.000 claims description 10
- 210000002751 lymph Anatomy 0.000 claims description 10
- 210000002700 urine Anatomy 0.000 claims description 10
- 241001148471 unidentified anaerobic bacterium Species 0.000 claims description 9
- 210000001215 vagina Anatomy 0.000 claims description 9
- 241000233866 Fungi Species 0.000 claims description 8
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 8
- DWAQJAXMDSEUJJ-UHFFFAOYSA-M Sodium bisulfite Chemical compound [Na+].OS([O-])=O DWAQJAXMDSEUJJ-UHFFFAOYSA-M 0.000 claims description 8
- 239000003795 chemical substances by application Substances 0.000 claims description 8
- 239000008103 glucose Substances 0.000 claims description 8
- 238000003756 stirring Methods 0.000 claims description 8
- 125000002791 glucosyl group Chemical group C1([C@H](O)[C@@H](O)[C@H](O)[C@H](O1)CO)* 0.000 claims description 5
- 230000000813 microbial effect Effects 0.000 claims description 5
- 239000007864 aqueous solution Substances 0.000 claims description 4
- 238000005238 degreasing Methods 0.000 claims description 4
- 238000001914 filtration Methods 0.000 claims description 4
- 239000003094 microcapsule Substances 0.000 claims description 4
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 claims description 3
- 229930091371 Fructose Natural products 0.000 claims description 3
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 claims description 3
- 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 3
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 claims description 3
- 150000002402 hexoses Chemical class 0.000 claims description 3
- 239000008101 lactose Substances 0.000 claims description 3
- LPHFLPKXBKBHRW-UHFFFAOYSA-L magnesium;hydrogen sulfite Chemical compound [Mg+2].OS([O-])=O.OS([O-])=O LPHFLPKXBKBHRW-UHFFFAOYSA-L 0.000 claims description 3
- JESHZQPNPCJVNG-UHFFFAOYSA-L magnesium;sulfite Chemical compound [Mg+2].[O-]S([O-])=O JESHZQPNPCJVNG-UHFFFAOYSA-L 0.000 claims description 3
- DJEHXEMURTVAOE-UHFFFAOYSA-M potassium bisulfite Chemical compound [K+].OS([O-])=O DJEHXEMURTVAOE-UHFFFAOYSA-M 0.000 claims description 3
- 229940099427 potassium bisulfite Drugs 0.000 claims description 3
- 235000010259 potassium hydrogen sulphite Nutrition 0.000 claims description 3
- RWPGFSMJFRPDDP-UHFFFAOYSA-L potassium metabisulfite Chemical compound [K+].[K+].[O-]S(=O)S([O-])(=O)=O RWPGFSMJFRPDDP-UHFFFAOYSA-L 0.000 claims description 3
- 229940043349 potassium metabisulfite Drugs 0.000 claims description 3
- 235000010263 potassium metabisulphite Nutrition 0.000 claims description 3
- 239000011734 sodium Substances 0.000 claims description 3
- 229910052708 sodium Inorganic materials 0.000 claims description 3
- 241000222120 Candida <Saccharomycetales> Species 0.000 claims description 2
- -1 malt carbohydrates Chemical class 0.000 claims description 2
- LSNNMFCWUKXFEE-UHFFFAOYSA-M Bisulfite Chemical compound OS([O-])=O LSNNMFCWUKXFEE-UHFFFAOYSA-M 0.000 claims 1
- 229940077731 carbohydrate nutrients Drugs 0.000 claims 1
- 229960002737 fructose Drugs 0.000 claims 1
- CXKWCBBOMKCUKX-UHFFFAOYSA-M methylene blue Chemical compound [Cl-].C1=CC(N(C)C)=CC2=[S+]C3=CC(N(C)C)=CC=C3N=C21 CXKWCBBOMKCUKX-UHFFFAOYSA-M 0.000 description 23
- 239000003086 colorant Substances 0.000 description 15
- 238000012360 testing method Methods 0.000 description 13
- 229960000583 acetic acid Drugs 0.000 description 11
- 239000000975 dye Substances 0.000 description 11
- 238000010186 staining Methods 0.000 description 11
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 9
- 230000028327 secretion Effects 0.000 description 9
- 235000010267 sodium hydrogen sulphite Nutrition 0.000 description 9
- 230000000052 comparative effect Effects 0.000 description 8
- 241000204031 Mycoplasma Species 0.000 description 7
- 229940079827 sodium hydrogen sulfite Drugs 0.000 description 7
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 6
- RAHZWNYVWXNFOC-UHFFFAOYSA-N Sulphur dioxide Chemical compound O=S=O RAHZWNYVWXNFOC-UHFFFAOYSA-N 0.000 description 6
- 238000007689 inspection Methods 0.000 description 6
- 150000007523 nucleic acids Chemical class 0.000 description 6
- 102000039446 nucleic acids Human genes 0.000 description 6
- 108020004707 nucleic acids Proteins 0.000 description 6
- 230000003647 oxidation Effects 0.000 description 6
- 238000007254 oxidation reaction Methods 0.000 description 6
- 230000008569 process Effects 0.000 description 6
- HRZFUMHJMZEROT-UHFFFAOYSA-L sodium disulfite Chemical compound [Na+].[Na+].[O-]S(=O)S([O-])(=O)=O HRZFUMHJMZEROT-UHFFFAOYSA-L 0.000 description 6
- 229940001584 sodium metabisulfite Drugs 0.000 description 6
- 235000010262 sodium metabisulphite Nutrition 0.000 description 6
- 201000008100 Vaginitis Diseases 0.000 description 5
- 229940095731 candida albicans Drugs 0.000 description 5
- 235000019441 ethanol Nutrition 0.000 description 5
- 238000000386 microscopy Methods 0.000 description 5
- ZCYVEMRRCGMTRW-UHFFFAOYSA-N 7553-56-2 Chemical compound [I] ZCYVEMRRCGMTRW-UHFFFAOYSA-N 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 4
- 206010046914 Vaginal infection Diseases 0.000 description 4
- 239000013078 crystal Substances 0.000 description 4
- 238000002845 discoloration Methods 0.000 description 4
- 150000004676 glycans Chemical class 0.000 description 4
- 229910052740 iodine Inorganic materials 0.000 description 4
- 239000011630 iodine Substances 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 244000000010 microbial pathogen Species 0.000 description 4
- 229920001282 polysaccharide Polymers 0.000 description 4
- 239000005017 polysaccharide Substances 0.000 description 4
- 230000035945 sensitivity Effects 0.000 description 4
- 238000012549 training Methods 0.000 description 4
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- MUBZPKHOEPUJKR-UHFFFAOYSA-N Oxalic acid Chemical compound OC(=O)C(O)=O MUBZPKHOEPUJKR-UHFFFAOYSA-N 0.000 description 3
- 239000002253 acid Substances 0.000 description 3
- 150000001412 amines Chemical class 0.000 description 3
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 238000003759 clinical diagnosis Methods 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- QTWZICCBKBYHDM-UHFFFAOYSA-N leucomethylene blue Chemical compound C1=C(N(C)C)C=C2SC3=CC(N(C)C)=CC=C3NC2=C1 QTWZICCBKBYHDM-UHFFFAOYSA-N 0.000 description 3
- 239000013642 negative control Substances 0.000 description 3
- 239000005715 Fructose Substances 0.000 description 2
- 208000007074 Trichomonas Vaginitis Diseases 0.000 description 2
- 208000025206 Trichomonas vaginitis urogenital infection Diseases 0.000 description 2
- 241000287436 Turdus merula Species 0.000 description 2
- 230000002378 acidificating effect Effects 0.000 description 2
- 150000001299 aldehydes Chemical class 0.000 description 2
- 230000001580 bacterial effect Effects 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 238000000546 chi-square test Methods 0.000 description 2
- 235000013399 edible fruits Nutrition 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 210000003743 erythrocyte Anatomy 0.000 description 2
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 2
- 229910052737 gold Inorganic materials 0.000 description 2
- 239000010931 gold Substances 0.000 description 2
- 230000002458 infectious effect Effects 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 150000007524 organic acids Chemical class 0.000 description 2
- 239000002504 physiological saline solution Substances 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 208000013464 vaginal disease Diseases 0.000 description 2
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 description 1
- ZNQVEEAIQZEUHB-UHFFFAOYSA-N 2-ethoxyethanol Chemical compound CCOCCO ZNQVEEAIQZEUHB-UHFFFAOYSA-N 0.000 description 1
- 229920002972 Acrylic fiber Polymers 0.000 description 1
- QGZKDVFQNNGYKY-UHFFFAOYSA-O Ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 241000207202 Gardnerella Species 0.000 description 1
- 206010061977 Genital infection female Diseases 0.000 description 1
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 238000003794 Gram staining Methods 0.000 description 1
- 102000004157 Hydrolases Human genes 0.000 description 1
- 108090000604 Hydrolases Proteins 0.000 description 1
- 206010021703 Indifference Diseases 0.000 description 1
- 244000283207 Indigofera tinctoria Species 0.000 description 1
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 102000005348 Neuraminidase Human genes 0.000 description 1
- 108010006232 Neuraminidase Proteins 0.000 description 1
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 description 1
- 208000030852 Parasitic disease Diseases 0.000 description 1
- SJEYSFABYSGQBG-UHFFFAOYSA-M Patent blue Chemical compound [Na+].C1=CC(N(CC)CC)=CC=C1C(C=1C(=CC(=CC=1)S([O-])(=O)=O)S([O-])(=O)=O)=C1C=CC(=[N+](CC)CC)C=C1 SJEYSFABYSGQBG-UHFFFAOYSA-M 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 description 1
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 0.000 description 1
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 1
- YHFXJKYHUWPWSJ-UHFFFAOYSA-L [Na+].[Na+].OS([O-])=O.OS([O-])=O Chemical group [Na+].[Na+].OS([O-])=O.OS([O-])=O YHFXJKYHUWPWSJ-UHFFFAOYSA-L 0.000 description 1
- 239000000980 acid dye Substances 0.000 description 1
- 229940126575 aminoglycoside Drugs 0.000 description 1
- 239000000908 ammonium hydroxide Substances 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000003125 aqueous solvent Substances 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 239000000981 basic dye Substances 0.000 description 1
- GUBGYTABKSRVRQ-QUYVBRFLSA-N beta-maltose Chemical compound OC[C@H]1O[C@H](O[C@H]2[C@H](O)[C@@H](O)[C@H](O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@@H]1O GUBGYTABKSRVRQ-QUYVBRFLSA-N 0.000 description 1
- 150000001719 carbohydrate derivatives Chemical class 0.000 description 1
- 239000003518 caustics Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 238000004042 decolorization Methods 0.000 description 1
- 238000005034 decoration Methods 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 150000002016 disaccharides Chemical class 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 210000002919 epithelial cell Anatomy 0.000 description 1
- 239000000796 flavoring agent Substances 0.000 description 1
- 235000019634 flavors Nutrition 0.000 description 1
- 239000012362 glacial acetic acid Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 150000002390 heteroarenes Chemical class 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 125000004435 hydrogen atom Chemical class [H]* 0.000 description 1
- 238000010166 immunofluorescence Methods 0.000 description 1
- 229910052738 indium Inorganic materials 0.000 description 1
- 208000027866 inflammatory disease Diseases 0.000 description 1
- 230000006651 lactation Effects 0.000 description 1
- 230000003458 metachromatic effect Effects 0.000 description 1
- MCPLVIGCWWTHFH-UHFFFAOYSA-L methyl blue Chemical compound [Na+].[Na+].C1=CC(S(=O)(=O)[O-])=CC=C1NC1=CC=C(C(=C2C=CC(C=C2)=[NH+]C=2C=CC(=CC=2)S([O-])(=O)=O)C=2C=CC(NC=3C=CC(=CC=3)S([O-])(=O)=O)=CC=2)C=C1 MCPLVIGCWWTHFH-UHFFFAOYSA-L 0.000 description 1
- 125000000325 methylidene group Chemical group [H]C([H])=* 0.000 description 1
- 230000002906 microbiologic effect Effects 0.000 description 1
- 150000002772 monosaccharides Chemical class 0.000 description 1
- 229910017604 nitric acid Inorganic materials 0.000 description 1
- 230000009965 odorless effect Effects 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 235000006408 oxalic acid Nutrition 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 230000003950 pathogenic mechanism Effects 0.000 description 1
- 230000007170 pathology Effects 0.000 description 1
- 230000000149 penetrating effect Effects 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- OARRHUQTFTUEOS-UHFFFAOYSA-N safranin Chemical compound [Cl-].C=12C=C(N)C(C)=CC2=NC2=CC(C)=C(N)C=C2[N+]=1C1=CC=CC=C1 OARRHUQTFTUEOS-UHFFFAOYSA-N 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 239000012047 saturated solution Substances 0.000 description 1
- 235000014347 soups Nutrition 0.000 description 1
- 238000004611 spectroscopical analysis Methods 0.000 description 1
- QAOWNCQODCNURD-UHFFFAOYSA-N sulfuric acid Substances OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 1
- 235000018553 tannin Nutrition 0.000 description 1
- 229920001864 tannin Polymers 0.000 description 1
- 239000001648 tannin Substances 0.000 description 1
- 235000012976 tarts Nutrition 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
- 239000000052 vinegar Substances 0.000 description 1
- 235000021419 vinegar Nutrition 0.000 description 1
- 210000002268 wool Anatomy 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/36—Gynecology or obstetrics
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/70—Mechanisms involved in disease identification
- G01N2800/7095—Inflammation
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Analytical Chemistry (AREA)
- Toxicology (AREA)
- Immunology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Physics & Mathematics (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
This application involves microorganism detection fields.Specifically, this application provides for detecting coloring composition of inflammatory lesion in the tissue of microorganism infection and subject and preparation method thereof, the kit for including this coloring composition is additionally provided.This coloring composition includes magenta, methylene blue, hydrochloric acid, carbohydrate reducing agent, acetic acid, water and any group or whole in being selected from the group：A) sodium sulfite and b) bisulfites and weighting sulphite.The coloring composition and kit of the application can be used for detecting the microorganism infection in microorganism infection, especially vaginal fluid and the inflammatory lesion in the tissue of subject.
Description
Technical field
The present invention relates to microorganism detection fields.Specifically, this application provides for detecting the dyeing of microorganism infection
Agent composition and preparation method thereof additionally provides the kit for including this coloring composition.The coloring agent combination of the application
Object and kit can be used for detecting the microorganism infection in microorganism infection, especially vaginal fluid.
Background technology
The inspection method of microorganism in microorganism, such as human vagina's secretion have cultivation, smear microscopy inspection,
Direct fluorescent immune method, biochemical enzyme process detection, immunology detection, detection of nucleic acids etc..Cultivation is to check trichomonad, candidiasis
The method for waiting pathogenic microorganisms most sensitive and special, but troublesome in poeration, time-consuming longer (5 days), it is difficult to as routine clinical inspection
Survey method.
It is micro- after microexamination secretion, dyeing after physiological saline and potassium hydroxide experiment that Smear microscopy, which is checked the mark,
Spectroscopy.Physiological saline is affected by temperature larger, the poor 60%-70 of sensibility of the method mainly for detection of trichomonad
%[1,2]；Potassium hydroxide is tested mainly for detection of candidiasis, but the poor 60%-80% of sensibility.It is shown after secretion dyeing
Micro mirror inspection, the method for dyeing have Gram's staining, Wright's staining, methylene blue staining.
Wright's staining is dyed using following solvent：Acid dyes：Yihong；Basic dye：Methylene blue；Dyestuff is molten
Agent：Methanol.Gram's staining is relatively good but relatively low for the detection sensitivity of trichomonad for candidiasis, the dyeing of bacterium class,
Because the characteristic unobvious of trichomonad after dyeing；Wright's staining and methylene blue staining advantage in terms of epithelial cell is observed are bright
It is aobvious, in terms of observation trichomonad and bacterium with Gram's staining indifference.The advantages of microexamination is specific high, but by subjectivity
Factor influences more.
The sensitivity and specificity of direct fluorescent immune method are in more than 80%-90%, but by the subjective shadow of diagnostic personnel
Sound is larger；Biochemical enzyme process detection has the detections such as PH detections, sialidase, Aminoglycoside enzyme, these Testing index are required to combine
Detection, sensibility and specificity are to be improved.
Detection of nucleic acids has trichomonad, candidiasis, a diplococcus detection of nucleic acids, more than 90% detection sensitivity, specifically
More than 90% property, the subjective impact by technical staff is smaller, but laboratory must meet the requirement in PCR laboratories.
More than vaginal fluid method of detecting pathogenic microorganism is laboratory examination, the time that clinical diagnosis must wait for
It is longer, it is unfavorable for clinical " look into and control " to vaginal inflammatory lesion, if cultivation needs 5 days or more, detection of nucleic acids needs 2
Hour or more, immunofluorescence technique needs 1 hour or more, and smear microscopy inspection and biochemical enzyme process are required to 30 minutes or more.Make
The microorganism in vaginal fluid is dyed with Wright Stain, whole process needs about half an hour, uses gram
Decoration method is also required at least about half an hour.
Therefore, this field is badly in need of a kind of microorganism in microorganism, such as vaginal fluid and passes through dyeing and color change
Change that time for being detected is short, accuracy rate is high, high specificity, coloring composition simple and efficient to handle.
Invention content
It was found by the inventors of the present invention that the sample from subject quickly can be detected to microbial staining using dyestuff
In microorganism, such as the microorganism in vaginal fluid.Basic principle is：Pinkish red and methylene blue is reduced to nothing first
Then color whereby it can be detected that specific microorganism using microbiological oxidation magenta and methylene blue colour developing.
The present invention provides a kind of for detecting the rapid dyeing agent composition of microorganism infection, to solve in the prior art
The existing above problem.The coloring composition of the application is particularly suitable for micro- life in the vaginal fluid of detection subject
Object.Sample is dyed with this rapid dyeing agent composition of the present invention, considerably improves the sensitivity of detection microorganism
Property and specificity, while easy to operate, in very short time, such as several seconds, at most no more than 1 minute in regard to can obtain as a result,
Required detection time is greatly shortened compared with the prior art, and is not required to technical training.
Therefore, the present invention provides a kind of coloring compositions, and it includes magenta, methylene blue, hydrochloric acid, carbohydrate reduction
Agent, acetic acid, water and any group or whole in being selected from the group：A) sodium sulfite and b) bisulfites and weighting sulfurous acid
Salt, wherein each component have following weight percent：
Wherein, it is zero when a) sodium sulfite and b) bisulfites are different with sulphite is laid particular stress on.
In one embodiment, in the coloring composition of the application, the content of sodium sulfite is 0-3%, preferably
For 0-2%, more preferably 0.3-1.5%, more preferably 0.3% or 1.5%.
In one embodiment, in the coloring composition of the application, the content of bisulfites is 0-2%, excellent
It is selected as 0-1.5%, more preferably 0.7-1.5%, more preferably 0.7% or 1.5%.
In one embodiment, in the coloring composition of the application, the content for laying particular stress on sulphite is 0-3%,
Preferably 0-1.5%, more preferably 1-1.5%, more preferably 1% or 1.5%.
In one embodiment, in the coloring composition of the application, pinkish red content is 0.05-1.5%, excellent
It is selected as 0.05-1.2%, more preferably 0.1-1.2%, more preferably 0.1% or 1.2%.
In one embodiment, in the coloring composition of the application, the content of methylene blue is 0.01-1%,
Preferably 0.05-0.9%, more preferably 0.2-0.9%, more preferably 0.2%, 0.5% or 0.9%.In an embodiment
In, in the coloring composition of the application, the content of methylene blue is 0.2-0.5%.In another embodiment, it is sub-
The content of methyl blue is 0.5-0.9%.
In one embodiment, in the coloring composition of the application, the content of hydrochloric acid is 0.1-3%, preferably
0.1-2%, preferably 0.6-1.6%, more preferably 0.6%, 1% or 1.6%.In one embodiment, the application's
In coloring composition, the content of hydrochloric acid is 0.6-1%.In another embodiment, the content of hydrochloric acid is 1-1.6%.
In a preferred embodiment, in the coloring composition of the application, bisulfites is selected from sulfurous acid
Hydrogen sodium, potassium bisulfite and magnesium bisulfite.
In a preferred embodiment, it in the coloring composition of the application, lays particular stress on sulphite and is selected from weighting
Sodium sulfite, potassium metabisulfite and weighting magnesium sulfite.
In one embodiment, in the coloring composition of the application, carbohydrate reducing agent be selected from glucose, fructose,
Galactolipin, hexose, lactose, malt carbohydrates and their derivative, more preferably content 0.1-2%, preferably 0.5-2%, 0.5-
1%, more preferably 0.5% or 1%.
In one embodiment, in the coloring composition of the application, the content of acetic acid is 0-6%, preferably 2-
5%, more preferably 2.9-4%, more preferably 2.9% or 4%.
In one embodiment, the pH of the coloring composition of the application be 3.8-5.4, preferably 4.7-5.2, more
Preferably 4.7-4.9 or 4.9-5.2.
Further aspect of the application is related to the method for preparing the coloring composition of the application, includes the following steps：
(a) magenta is dissolved in distilled water, filtering forms solution 1, and the distilled water is preferably 80 DEG C of distilled water；
(b) by a) sodium sulfite and b), any one of bisulfites and weighting sulphite or whole aqueous solutions add
Enter and stirred in the solution 1 obtained in (a), obtain solution 2；
(c) hydrochloric acid is added in the solution 2 obtained in (b) to stir to pale red, add distilled water, obtain solution 3；
(d) methylene blue is added in stirring in the solution 3 obtained in (c) and, to dissolving, obtains solution 4；
(e) acetic acid is added in the solution 4 obtained in (d) and stirred, obtain solution 5；
(f) carbohydrate reducing agent is added in the solution 5 obtained in (e), stirring obtains the coloring agent combination to dissolving
Object.
Above steps all carries out under normal pressure.
In one aspect, coloring composition according to the present invention or coloring agent prepared according to the methods of the invention combination
The inflammatory lesion that object can be used in the tissue of the microorganism infection in sample of the detection from subject or subject, wherein described
Tissue is preferably vagina or skin etc., and the sample is preferably vaginal fluid, urine, lymph or biopsy etc.,
And the subject is preferably mammal, more preferably people, monkey, goat, pig or horse etc..
In one aspect, it is micro- in the sample from subject is detected this application involves the coloring composition of the application
Purposes in biological infection, the sample are preferably vaginal fluid, urine, lymph or biopsy etc., and described
Subject is preferably mammal, more preferably people, monkey, goat, pig or horse etc..
In yet another aspect, present invention also provides the coloring compositions comprising the application and the optionally examination of carrier
Agent box.In a preferred embodiment, this kit is for the microorganism infection in sample of the detection from subject.
Preferably, sample is vaginal fluid, urine, lymph or biopsy etc..On the other hand, subject is preferably lactation
Animal, more preferably people, monkey, goat, pig or horse etc..
In a preferred embodiment, carrier includes but not limited to degreasing cotton swab, gauze, micro-capsule, tunica fibrosa, filter paper
Deng.
The application further relates to purposes of the kit in the microorganism infection in detecting the sample from subject, the sample
Product are preferably vaginal fluid, urine, lymph or biopsy etc., and the subject is preferably mammal, more
Preferably people, monkey, goat, pig or horse etc., the microorganism are preferably fungi, such as candidiasis, trichomonad, anaerobic bacteria, mould
Bacterium or Gardner bacillus etc..
Gardner bacillus is also known as Hemophilus vaginalis(Hemophilus vaginalis), belongs to Gardnerella, the entitled vagina Gartner bacterium of bacterium, (G
Vaginalis), referred to as GV.Gardner bacillus belongs to anaerobic bacteria.
The invention further relates to detection subject tissue in inflammatory lesion method, the method includes：
(a) sample from the tissue is acquired using carrier, and uses the coloring composition or reagent of the application
Coloring composition in box is to the carrier dyeing；
(b) color change on carrier is observed；
(c) according to color change determine subject tissue whether microbial infection.
In a preferred embodiment, it is of the present invention detection subject tissue in inflammatory lesion method
In, the observation in step (b) is carried out by naked eyes.
In a preferred embodiment, the microorganism that the coloring composition of the application or kit are detected is true
Bacterium (such as candidiasis), trichomonad, anaerobic bacteria, mould or Gardner bacillus etc..
In a preferred embodiment, it is of the present invention detection subject tissue in inflammatory lesion method
In, it determines to be carried out according to following standard in the observation and the step (c) in the step (b)：Carrier does not change colour, prompts
Tissue is without inflammatory lesion；Carrier changes colour, and there are inflammatory lesions in prompting tissue.
The coloring composition of the different modes application present invention can be passed through.In one embodiment, by using institute
Coloring composition is stated to dye the carrier for acquiring the sample from subject.In a preferred embodiment, lead to
Cross the sample that the coloring composition in the coloring composition or the kit is applied to the subject by carrier smearing
Product.Preferably, the carrier includes but not limited to degreasing cotton swab, gauze, micro-capsule, tunica fibrosa, filter paper etc..
In one embodiment, wherein the tissue is preferably vagina or skin etc., the sample is preferably vagina point
Secretion, urine, lymph or biopsy etc..In another embodiment, the subject is preferably mammal,
More preferably people, monkey, goat, pig or horse etc..
The invention further relates to the application coloring composition or kit detection subject tissue in inflammatory disease
Purposes in change.In one embodiment, tissue is preferably vagina or skin etc..In another embodiment, subject
Preferably mammal, more preferably people, monkey, goat, pig or horse etc..
Degree involved in the present invention is weight percentage, unless otherwise specified.
Attached drawing describes
The result dyed with the coloring composition of the application to vaginal fluid is shown in Fig. 1.A in Fig. 1
Negative in embodiment 1,2 and 3, infectious result is shown.B in Fig. 1 is shown in embodiment 1,2 and 3 that there are micro-
The colour developing result of biological infection.By, it is found that carrier does not change colour, it is to be uninfected by micro- life that yellow is shown as on colorimetric card in Fig. 1
The normal healthy controls (feminine gender) of object.In contrast, carrier changes colour, for example, being shown as blue, green or celadon are the micro- life of infection
The positive patient of object.
Specific embodiment
The coloring composition and/or can quickly detect microorganism infection comprising its kit that the application provides, it is special
Be not subject vaginal fluid in microorganism infection.The coloring composition of the application can obtain dyeing knot in real time
Fruit, i.e., after sample has been acquired, coloration result can come out.Without waiting for the processing such as film-making, the dyeing in laboratory.
Sample is dyed with this rapid dyeing agent composition of the present invention, considerably improves detection microorganism
Sensibility and specificity, while easy to operate, in very short time, such as several seconds, at most no more than 1 minute in regard to that can be tied
Fruit, required detection time is greatly shortened compared with the prior art, and is not required to technical training.
Specifically, in the coloring composition of the application, in the pinkish red acidic environment provided in hydrochloric acid with sodium sulfite
Or bisulfites and weighting sulphite reaction are reduced to colourless pinkish red (Schiff's reagent), methylene blue is by carbohydrate reducing agent
It is reduced to leucomethylene blue.When being dyed using vaginal fluid coloring agent, the microorganism in secretion can make colourless
Pinkish red or leucomethylene blue occurs oxidation and develops the color, and the color of coloring composition as a result can be made to change, by no discoloration
Into other colors, so as to be used to detect microorganism.
The aldehyde radical for example, polysaccharide of fungi (such as candidiasis, mould) bacterium wall in secretion dissociates, the aldehyde radical with
Colourless magenta is developed the color (magenta or purple) with reference to the new pinkish red compound of generation, whereby it can be detected that fungi；Meanwhile
The thermophilic methylene blue of protease such as acid hydrolase of trichomonad secretion and aobvious blue, so as to detect trichomonad；Also, anaerobic bacteria,
Such as a large amount of active oxygen such as H that Gardner bacillus generates2O2It aoxidizes leucomethylene blue and aobvious green, whereby it can be detected that
Anaerobic bacteria, it is thus achieved that of the invention.
Therefore, when the coloring composition of the application is used to detect the microorganism in microorganism, such as tissue, can make
The color of coloring composition changes, and becomes other colors by colourless, so as to be used to detect microorganism.
Magenta is a kind of common coloring agent, for the dyeing of silk, acrylic fibers, wool and tannin mordant dyeing cotton fiber, is encountered
Sulfur dioxide solution decolourization.Dark red powder.Water is dissolved in blue light red to magenta, is slightly soluble in ethyl alcohol, acetone and Cellosolve,
Insoluble in other organic solvents.It is red in blue light to meet the concentrated sulfuric acid, in shiny red after dilution, meets concentrated nitric acid to turn after blush solution
It is orange.Pinkish red aqueous solution enriching hydrochloric acid adds caustic lye of soda in orange-brown in magenta.In the coloring composition of the application
And preparation method thereof in pinkish red content be 0.05-1.5%, more preferably preferably 0.05-1.2%, 0.1-1.2%, it is more excellent
It is selected as 0.1% or 1.2%.
Methylene blue is a kind of clinically common coloring agent.It is a kind of heteroaromatic compound.It is used as chemistry to refer to
Show that agent, dyestuff, biological stain and drug use.Methylene blue staining, metachromatic principle are mainly based upon it in oxidation state and go back
Different colours in ortho states.Specifically, the aqueous solution of methylene blue is aoxidized in oxidative environment, i.e. the methylene of oxidation state
Base indigo plant in blue, but meet zinc, the reducing agents such as ammonium hydroxide can be reduced, i.e. the methylene blue of reduction-state is into colourless form.In the application
The content of coloring composition and preparation method thereof Methylene Blue be 0.01-1%, preferably 0.05-0.9%, more preferably
For 0.2-0.9%, more preferably 0.2%, 0.5% or 0.9%.In one embodiment, it is combined in the coloring agent of the application
In object, the content of methylene blue is 0.2-0.5%.In another embodiment, the content of methylene blue is 0.5-0.9%.
The hydrochloric acid used in coloring composition of the present invention and preparation method thereof provides acid environment, preferably
0.1-2%.In coloring composition of the application and preparation method thereof, the content of hydrochloric acid is 0.1-3%, preferably 0.1-
2%, preferably 0.6-1.6%, more preferably 0.6%, 1% or 1.6%.In one embodiment, in the dyeing of the application
In agent composition, the content of hydrochloric acid is 0.6-1%.In another embodiment, the content of hydrochloric acid is 1-1.6%.
Hydrochloric acid is the constituent of Schiff's reagent in the coloring composition of the application, provides acidic environment.
The pH of the coloring composition of the application be 3.8-5.4, preferably 4.7-5.2, more preferably 4.7-4.9 or
4.9-5.2。
" carbohydrate reducing agent " in the present invention refers to any carbohydrate with reproducibility, its derivative and combinations thereof.The sugar
Class includes but not limited to monosaccharide, disaccharides, polysaccharide.Specifically, the carbohydrate reducing agent can be glucose, fructose, galactolipin,
Hexose, lactose, maltose etc.." carbohydrate derivative " described herein is defined as glucide by polymerizeing, being esterified, aoxidizing
React such as polysaccharide formed, glycoprotein, organic acid derivative.Coloring composition and preparation of the carbohydrate reducing agent in the present invention
Content in method is 0.1-2%, more preferably preferably 0.5-2%, 0.5-1%, more preferably 0.5% or 1%.
Oxidation state methylene blue is reduced to by the carbohydrate reducing agent in coloring composition of the present invention and preparation method thereof
Colourless reduction-state methylene blue.
Sodium sulfite in coloring composition of the application and preparation method thereof is used to magenta being reduced to no chromaticity
It is red.The amount of sodium sulfite in coloring composition of the application and preparation method thereof be 0-3%, preferably 0-2%, more
Preferably 0.3-1.5%, more preferably 0.3% or 1.5%.
" bisulfites " loses part sulfur dioxide in exposure air in the application, and simultaneous oxidation is into sulfate.
Bisulfites in coloring composition of the application and preparation method thereof is also used for magenta being reduced to colourless magenta.At this
The content of bisulfites in coloring composition of application and preparation method thereof be 0-2%, preferably 0-1.5%, it is more excellent
It is selected as 0.7-1.5%, more preferably 0.7% or 1.5%.In a preferred embodiment, in the coloring agent group of the application
It closes in object and method, the bisulfites is selected from sodium hydrogensulfite, potassium bisulfite and magnesium bisulfite.
" sulphite is laid particular stress in the application " to contact with strong acid, is released sulfur dioxide and is generated corresponding salt, is long placed in
It can be aoxidized in air.Weighting sulphite in coloring composition in the application and preparation method thereof is also used for product
It is red to be reduced to colourless magenta.The content of weighting sulphite in coloring composition of the application and preparation method thereof is 0-
3%, preferably 0-1.5%, more preferably 1-1.5%, more preferably 1% or 1.5%.In a preferred embodiment,
In the coloring composition and method of the application, lay particular stress on sulphite be selected from Sodium Metabisulfite, potassium metabisulfite and partially
Weight magnesium sulfite etc..
Sodium sulphite, bisulfites, can magenta be reduced to colourless magenta by laying particular stress on any one of sulphite.Nothing
Chromaticity is red to act on aobvious purple with aldehyde, does not develop the color with ketolysis.This chromogenic reaction is very sensitive, available for differentiating aldehydes chemical combination
Object, such as the polysaccharide of fungi (mainly candidiasis) bacterium wall dissociate the aldehyde radical, so as to be used to identify Candida
Bacterium.Magenta is reduced to no chromaticity by the combination of sodium sulfite or bisulfites used herein and weighting sulphite
It is red.
In the application " acetic acid ", acetic acid, glacial acetic acid, chemical formula CH are also3COOH is a kind of organic acid, is vinegar
The source of interior tart flavour and penetrating odor.There may be red blood cells in vaginal fluid, and in of the invention, the effect of acetic acid is to reduce
Influence of the red blood cell to color interpretation.In coloring composition of the application and preparation method thereof, the content of acetic acid is 0-
6%, preferably 2-5%, preferably 2.9-4%, more preferably 2.9% or 4%.
" water " (H in the application2O it is) a kind of common solvent, is at normal temperatures and pressures the transparency liquid of colorless and odorless.
In coloring composition of the application and preparation method thereof, the content of water is 78.5-96%.
It is further illustrated the present invention below by embodiment and comparative example.In the following Examples and Comparative Examples, with nucleic acid
Testing result is goldstandard, and the recall rate of coloring agent is illustrated with sensibility, and sensibility is higher, finds the ability of pathogenic microorganisms
By force；Illustrate the accuracy of coloring agent with specificity, specificity is higher, and the pathogenic microorganisms and detection of nucleic acids result for showing detection accord with
Conjunction rate is high.
In one embodiment, in the coloring composition of the application, bisulfites is sodium hydrogensulfite, is laid particular stress on
Bisulfites is Sodium Metabisulfite, and carbohydrate reducing agent is glucose.
In one embodiment, it in the coloring composition of the application, is gone back using sodium sulfite as reducing agent
Original is red, and wherein the content of sodium sulfite is 1.5%, and pinkish red content is 0.1%, and the content of methylene blue is 0.2%, hydrochloric acid
Content for 1.6%, carbohydrate reducing agent is glucose, and content 1.0%, the content of acetic acid is 4.0%, coloring composition
PH be 4.7.
In one embodiment, in the coloring composition of the application, while sodium sulfite, sodium hydrogensulfite are used
Magenta is restored as reducing agent, wherein the content of sodium sulfite is 0.3%, sodium hydrogensulfite contains with Sodium Metabisulfite
It is 0.7% to measure, and the content of Sodium Metabisulfite is 1.5%, and pinkish red content is 0.1%, and the content of methylene blue is 0.5%,
The content of hydrochloric acid is 1.0%, and carbohydrate reducing agent is glucose, and content 0.5%, the content of acetic acid is 4.0%, coloring agent group
The pH for closing object is 4.9.
In one embodiment, in the coloring composition of the application, using sodium hydrogensulfite and sulfurous acid is laid particular stress on
Sodium restores magenta as reducing agent, and the wherein content of sodium hydrogensulfite is 1.5%, and the content of Sodium Metabisulfite is 1.0%,
Pinkish red content is 1.2%, and the content of methylene blue is 0.9%, and the content of hydrochloric acid is 0.6%, and carbohydrate reducing agent is glucose,
Its content is 0.5%, and the content of acetic acid is 2.9%, and the pH of coloring composition is 5.2.
In the present invention, sensibility and specificity is defined as below and calculates：
Sensibility=true positives number/(true positives number+false negative number) * 100%
Specificity=true negative number/(true negative number+false positive number) * 100%.
Embodiment
The patient that 80 Out-patient Clinic of Department of Gynecology are gone to a doctor is randomly selected, vaginal fluid is acquired with medical cotton swab, carries out embodiment
1st, 2,3 Coloration experiment and the microscopy of comparative example are carried out at the same time amine test, test pH and Culture Mycoplasma, trichomonad culture
It is cultivated with candida albicans, specific method is as follows：
Embodiment 1：
At ambient temperature and pressure, after magenta is dissolved in 80 DEG C of distilled water by each component specified in following table 1 respectively, cooling,
Sodium sulfite, Sodium Metabisulfite stirring are added in after filtering, hydrochloric acid is added in and stirs to pale red, add distilled water, is added in sub-
Methyl blue and glucose are stirred to dissolving, and rear addition acetic acid, which is stirred, is made coloring composition.Use coloring composition
To acquire vaginal fluid cotton swab dye, observe the color change of cotton swab, if cotton swab without discoloration, for feminine gender, table
Show no bacterium or microorganism infection；If cotton swab changes colour (such as shown in green or blue), for the positive, prompt have vagina
Inflammation.
Amine test is carried out to sample simultaneously, pH is tested and Culture Mycoplasma, trichomonad cultivate and candida albicans culture, and with
Using Amsel standard laws as goldstandard come diagnosing bacterial vagina disease (BV), Culture Mycoplasma is positive, trichomonad culture is positive and
The positive gold mark as clinical diagnosis mycoplasma vaginitis, trichomonas vaginitis and candidal vaginitis of candida albicans culture
It is accurate[6]-[12], the sensibility and specificity of coloring composition is counted, test result is listed in table 1.Detect trichomonad,
The infection of the microorganisms such as mould, Gardner bacillus.Negative control is illustratively given in Fig. 1 and the part of positive is compared
As a result.
Embodiment 2
The method of embodiment 1 is repeated by each component content specified in following table 1, but sulfurous acid is replaced with sodium hydrogensulfite
Sodium lists test result in table 1.Detect the infection of the microorganisms such as trichomonad, mould, Gardner bacillus.Example in Fig. 1
Give negative control and the part comparison result of positive to property.As shown in Figure 1, carrier does not change colour, in colorimetric
It is to be uninfected by the normal healthy controls (feminine gender) of microorganism that yellow is shown as on card, and carrier discoloration is shown as blue, green or grayish green
Color is the positive patient of microbial infection.Detect the infection of the microorganisms such as trichomonad, mould, Gardner bacillus.
Embodiment 3
The method that embodiment 1 is repeated by each component content specified in following table 1, but sodium sulfite and sodium hydrogensulfite are total to
With being dissolved in aqueous solvent, suffered in table 1 and listed test result.Negative control and the positive are illustratively given in Fig. 1
The part comparison result of sample.
Meanwhile illustratively show vaginal fluid is dyed with the coloring composition of the application in Fig. 1
As a result.Negative in experiment three times, infectious result is shown in A in Fig. 1.B in Fig. 1 is shown that there are microorganism senses
The colour developing result of dye.As shown in Figure 1, carrier does not change colour, and it is to be uninfected by microorganism that yellow is shown as on colorimetric card
Normal healthy controls (feminine gender), carrier discoloration is shown as blue, green or positive patient that celadon is microbial infection.Inspection
The infection of the microorganisms such as trichomonad, mould, Gardner bacillus is measured.
Embodiment 1-3 only needs about 30 seconds with regard to that can obtain result.
Comparative example
The microorganism in vaginal fluid is dyed using Gram's stain, whole process needs about half small
When.Specifically, at ambient temperature and pressure, gram staining liquid is configured：By crystal violet ethyl alcohol saturated solution, (crystal violet 2g is dissolved in
In 20mL95% ethyl alcohol) 20mL and 1% oxalic acid aqueous ammonium 80mL mixings put for 24 hours that filtering is made into crystal violet liquid afterwards；By 0.66g iodine
Change potassium to be dissolved in a small amount of distilled water, then add in 0.33g iodine and be allowed to be completely dissolved, add distilled water to 100mL and be made into Lu Ge
(Lugol) family name's iodine solution；Sarranine 0 (safranine, also known as husky of common dye 0) 0.25g is dissolved in 95% ethyl alcohol of 10mL, then is steamed with 40ml
Distilled water mixing is made into sarranine solution.After secretion is made smear drying, crystal violet liquid is added to be washed after dyeing 1 minute, Jia Luge
Family name's iodine staining is washed after 1 minute, and 95% alcohol is added to be washed after decolourizing 40 seconds, sucks moisture, in addition sarranine solution dyes 1 point
Clock is washed, and dries rear microscopy in air, and whole process needs at least about half an hour.
Amine test is carried out to vaginal fluid simultaneously, tests pH and Culture Mycoplasma, trichomonad culture and candida albicans training
It supports, and to be trained using Amsel standard laws as goldstandard come diagnosing bacterial vagina disease (BV), the Culture Mycoplasma positive, trichomonad
Establishing the yang function property and the positive gold as clinical diagnosis mycoplasma vaginitis, trichomonas vaginitis and candidal vaginitis of candida albicans culture
Standard tests Gram-stained sensibility and specificity, and test result is listed in table 1.Also, come using Chi-square Test
Verification embodiment 1-3 is respectively relative to the conspicuousness of the difference of the result of comparative example.
Table 1
Remarks：＆, *, # represent p<0.01 (Chi-square Test), i.e. data have significant significant difference
The sensibility of embodiment and comparative example coloring agent, the test data explanation of specificity, the moon of the invention in above-mentioned table 1
Road secretion coloring composition specificity is suitable with comparative example, but the sensibility in embodiment 1-3 will be significantly higher than comparative example.
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Claims (25)
1. coloring composition, it includes magenta, methylene blue, hydrochloric acid, carbohydrate reducing agent, acetic acid, water and it is selected from the group
Any group or whole：A) sodium sulfite and b) bisulfites and weighting sulphite, wherein each component is with following weight hundred
Divide ratio：
Wherein, it is zero when a) sodium sulfite and b) bisulfites are different with sulphite is laid particular stress on.
2. the content of coloring composition described in claim 1, wherein sodium sulfite is 0-2%, preferably 0.3-1.5%,
More preferably 0.3% or 1.5%.
3. the coloring composition described in claims 1 or 2, the content of bisulfite is 0-1.5%, preferably 0.7-
1.5%, more preferably 0.7% or 1.5%.
4. the coloring composition described in any one of claim 1-3, wherein the content for laying particular stress on sulphite is 0-1.5%,
Preferably 1-1.5%, more preferably 1% or 1.5%.
5. the coloring composition described in any one of preceding claims, wherein pinkish red content is 0.05-1.2%, preferably
For 0.1-1.2%, more preferably 0.1% or 1.2%.
6. the coloring composition described in any one of preceding claims, the content of Methylene Blue is 0.05-0.9%,
Preferably 0.2-0.9%, preferably 0.2-0.5% or preferably 0.5-0.9%, more preferably 0.2%, 0.5% or 0.9%.
7. the content of the coloring composition described in any one of preceding claims, wherein hydrochloric acid is 0.1-2%, preferably
0.6-1.6%, preferably 0.6-1% or preferably 1-1.6%, more preferably 0.6%, 1% or 1.6%.
8. the coloring composition described in any one of preceding claims, wherein the bisulfites is selected from bisulfite
Sodium, potassium bisulfite and magnesium bisulfite.
9. the coloring composition described in any one of preceding claims, wherein the weighting sulphite is selected from and lays particular stress on sulfurous
Sour sodium, potassium metabisulfite and weighting magnesium sulfite.
10. the coloring composition described in any one of preceding claims, wherein the carbohydrate reducing agent is selected from glucose, fruit
Sugar, galactolipin, hexose, lactose, malt carbohydrates and their derivative, content are preferably 0.5-2%, more preferably 0.5-1%, more preferably
It is 0.5% or 1%.
11. the content of the coloring composition described in any one of preceding claims, wherein acetic acid is 2-5%, preferably
2.9-4%, more preferably 2.9% or 4%.
12. the coloring composition described in any one of preceding claims, wherein the pH of the coloring composition is 3.8-
5.4, preferably 4.7-5.2, more preferably 4.7-4.9 or 4.9-5.2.
13. the coloring composition described in any one of preceding claims, wherein the coloring composition comes for detecting
From the microorganism infection in the sample of subject or the inflammatory lesion in the tissue of subject, it is described tissue be preferably vagina or
Skin etc., the sample is preferably vaginal fluid, urine, lymph or biopsy etc., and the subject is preferred
It is mammal, more preferably people, monkey, goat, pig or horse etc., the microorganism are preferably fungi, such as candidiasis,
Trichomonad, anaerobic bacteria, mould or Gardner bacillus etc..
14. microorganism of the coloring composition in the sample from subject is detected described in any one of preceding claims
Purposes in infection, the sample are preferably vaginal fluid, urine, lymph or biopsy etc., and described tested
Person is preferably mammal, more preferably people, monkey, goat, pig or horse etc., and the microorganism is preferably fungi, such as false silk ferment
Female bacterium, trichomonad, anaerobic bacteria, mould or Gardner bacillus etc..
15. preparing the method for the coloring composition described in any one of claim 1-13, include the following steps：
(a) magenta is dissolved in distilled water, filtering forms solution 1, and the distilled water is preferably 80 DEG C of distilled water；
(b) by a) sodium sulfite and b), any one of bisulfites and weighting sulphite or whole aqueous solutions are added to
(a) it is stirred in the solution 1 obtained in, obtains solution 2；
(c) hydrochloric acid is added in the solution 2 obtained in (b) to stir to pale red, add distilled water, obtain solution 3；
(d) methylene blue is added in stirring in the solution 3 obtained in (c) and, to dissolving, obtains solution 4；
(e) acetic acid is added in the solution 4 obtained in (d) and stirred, obtain solution 5；
(f) carbohydrate reducing agent is added in the solution 5 obtained in (e), stirring obtains the coloring composition to dissolving；
Above steps all carries out under normal pressure.
16. kit, it includes the coloring composition described in any one of claim 1-13 and optionally carrier, preferably
The ground kit for detect the sample from subject in microorganism infection, the sample be preferably vaginal fluid,
Urine, lymph or biopsy etc., and the subject is preferably mammal, more preferably people, monkey, goat,
Pig or horse etc., the microorganism are preferably fungi, such as candidiasis, trichomonad, anaerobic bacteria, mould or Gardner bacillus
Deng.
17. the kit described in claim 16, wherein the carrier be selected from degreasing cotton swab, gauze, micro-capsule, tunica fibrosa or
Filter paper.
18. the method for inflammatory lesion in the tissue of subject is detected, the method includes：
(a) sample from the tissue, and the dyeing described in using any one of claim 1-13 are acquired using carrier
Coloring composition in the kit of any one of agent composition or claim 16-17 is to the carrier dyeing；
(b) color change on carrier is observed；
(c) according to color change determine subject tissue whether microbial infection.
19. the method for claim 18, wherein the observation in the step (b) is carried out by naked eyes.
20. the method for claim 18 or 19, wherein the determining root in observation and the step (c) in the step (b)
Standard is descended to carry out according to this：Carrier does not change colour, prompts the tissue without inflammatory lesion；Carrier changes colour, and prompts to exist in the tissue
Inflammatory lesion.
21. the method described in any one of claim 18-20, wherein the carrier include but not limited to degreasing cotton swab, gauze,
Micro-capsule, tunica fibrosa or filter paper.
22. the method described in any one of claim 18-20, wherein being smeared by carrier by the coloring composition or institute
State the sample that the coloring composition in kit is applied to the subject.
23. the method described in any one of claim 18-22, wherein the tissue is preferably vagina or skin etc., the sample
Product are preferably vaginal fluid, urine, lymph or biopsy etc., and the subject is preferably mammal, more
Preferably people, monkey, goat, pig or horse etc..
24. the examination described in any one of coloring composition or claim 16-17 described in any one of claim 1-13
Purposes in inflammatory lesion of the agent box in the tissue of detection subject, the tissue are preferably vagina or skin etc., and institute
It states subject and is preferably mammal, more preferably people, monkey, goat, pig or horse etc..
25. the kit described in any one of claim 16-17 is in the microorganism sense in the sample from subject is detected
Purposes in dye, the sample are preferably vaginal fluid, urine, lymph or biopsy etc., and the subject
Preferably mammal, more preferably people, monkey, goat, pig or horse etc., the microorganism is preferably fungi, such as Candida
Bacterium, trichomonad, anaerobic bacteria, mould or Gardner bacillus etc..
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810048667.7A CN108148887A (en) | 2018-01-18 | 2018-01-18 | Coloring composition for detecting microorganism infection and preparation method thereof |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810048667.7A CN108148887A (en) | 2018-01-18 | 2018-01-18 | Coloring composition for detecting microorganism infection and preparation method thereof |
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| CN108148887A true CN108148887A (en) | 2018-06-12 |
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| CN201810048667.7A Pending CN108148887A (en) | 2018-01-18 | 2018-01-18 | Coloring composition for detecting microorganism infection and preparation method thereof |
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