EP0051593A1 - Isolation of carbohydrates from cell tissue - Google Patents
Isolation of carbohydrates from cell tissueInfo
- Publication number
- EP0051593A1 EP0051593A1 EP81900324A EP81900324A EP0051593A1 EP 0051593 A1 EP0051593 A1 EP 0051593A1 EP 81900324 A EP81900324 A EP 81900324A EP 81900324 A EP81900324 A EP 81900324A EP 0051593 A1 EP0051593 A1 EP 0051593A1
- Authority
- EP
- European Patent Office
- Prior art keywords
- galp
- process according
- glc
- gal
- blood
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Withdrawn
Links
- 150000001720 carbohydrates Chemical class 0.000 title claims abstract description 21
- 235000014633 carbohydrates Nutrition 0.000 title claims abstract description 11
- 238000002955 isolation Methods 0.000 title description 5
- 150000002482 oligosaccharides Chemical class 0.000 claims abstract description 44
- 229920001542 oligosaccharide Polymers 0.000 claims abstract description 42
- 239000008280 blood Substances 0.000 claims abstract description 37
- 238000000034 method Methods 0.000 claims abstract description 30
- 210000004369 blood Anatomy 0.000 claims abstract description 29
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical class OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 claims abstract description 11
- QAEDZJGFFMLHHQ-UHFFFAOYSA-N trifluoroacetic anhydride Chemical compound FC(F)(F)C(=O)OC(=O)C(F)(F)F QAEDZJGFFMLHHQ-UHFFFAOYSA-N 0.000 claims abstract description 8
- 238000006243 chemical reaction Methods 0.000 claims abstract description 5
- 230000015556 catabolic process Effects 0.000 claims abstract description 4
- 238000006731 degradation reaction Methods 0.000 claims abstract description 4
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract 5
- 230000008569 process Effects 0.000 claims description 18
- 239000012528 membrane Substances 0.000 claims description 16
- 239000000376 reactant Substances 0.000 claims description 9
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 7
- 239000011541 reaction mixture Substances 0.000 claims description 5
- 239000002904 solvent Substances 0.000 claims description 5
- 238000005227 gel permeation chromatography Methods 0.000 claims description 3
- 238000004816 paper chromatography Methods 0.000 claims description 3
- 239000012071 phase Substances 0.000 claims description 3
- 239000008346 aqueous phase Substances 0.000 claims description 2
- 210000004027 cell Anatomy 0.000 abstract description 14
- 210000003617 erythrocyte membrane Anatomy 0.000 abstract 1
- 210000001519 tissue Anatomy 0.000 description 12
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 10
- 210000004379 membrane Anatomy 0.000 description 10
- 229930186217 Glycolipid Natural products 0.000 description 6
- 238000002360 preparation method Methods 0.000 description 6
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 5
- 102000003886 Glycoproteins Human genes 0.000 description 5
- 108090000288 Glycoproteins Proteins 0.000 description 5
- 241001465754 Metazoa Species 0.000 description 5
- 230000008878 coupling Effects 0.000 description 4
- 238000010168 coupling process Methods 0.000 description 4
- 238000005859 coupling reaction Methods 0.000 description 4
- 235000018102 proteins Nutrition 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- 102000004169 proteins and genes Human genes 0.000 description 4
- 238000000746 purification Methods 0.000 description 4
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- HUMHYXGDUOGHTG-HEZXSMHISA-N alpha-D-GalpNAc-(1->3)-[alpha-L-Fucp-(1->2)]-D-Galp Chemical compound O[C@H]1[C@H](O)[C@H](O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](O[C@@H]2[C@@H]([C@@H](O)[C@@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](O)[C@@H](CO)OC1O HUMHYXGDUOGHTG-HEZXSMHISA-N 0.000 description 3
- 239000000969 carrier Substances 0.000 description 3
- 238000007385 chemical modification Methods 0.000 description 3
- 150000002298 globosides Chemical class 0.000 description 3
- 238000002649 immunization Methods 0.000 description 3
- 239000003547 immunosorbent Substances 0.000 description 3
- 150000002632 lipids Chemical class 0.000 description 3
- WQZGKKKJIJFFOK-SVZMEOIVSA-N (+)-Galactose Chemical compound OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-SVZMEOIVSA-N 0.000 description 2
- -1 2-acetamido-2-deoxy-D-galactopyranosyl Chemical group 0.000 description 2
- 125000003047 N-acetyl group Chemical group 0.000 description 2
- 229920005654 Sephadex Polymers 0.000 description 2
- 239000012507 Sephadex™ Substances 0.000 description 2
- 125000003277 amino group Chemical group 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
- 239000012876 carrier material Substances 0.000 description 2
- 210000000170 cell membrane Anatomy 0.000 description 2
- 210000000981 epithelium Anatomy 0.000 description 2
- 230000006870 function Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- DCXYFEDJOCDNAF-UHFFFAOYSA-N Asparagine Natural products OC(=O)C(N)CC(N)=O DCXYFEDJOCDNAF-UHFFFAOYSA-N 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 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 description 1
- LCWXJXMHJVIJFK-UHFFFAOYSA-N Hydroxylysine Natural products NCC(O)CC(N)CC(O)=O LCWXJXMHJVIJFK-UHFFFAOYSA-N 0.000 description 1
- 238000006994 Koenigs-Knorr glycosidation reaction Methods 0.000 description 1
- SHZGCJCMOBCMKK-DHVFOXMCSA-N L-fucopyranose Chemical compound C[C@@H]1OC(O)[C@@H](O)[C@H](O)[C@@H]1O SHZGCJCMOBCMKK-DHVFOXMCSA-N 0.000 description 1
- PNNNRSAQSRJVSB-UHFFFAOYSA-N L-rhamnose Natural products CC(O)C(O)C(O)C(O)C=O PNNNRSAQSRJVSB-UHFFFAOYSA-N 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 230000021736 acetylation Effects 0.000 description 1
- 238000006640 acetylation reaction Methods 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 125000001931 aliphatic group Chemical group 0.000 description 1
- XNBZPOHDTUWNMW-OUUCXATCSA-N alpha-L-Fucp-(1->2)-[alpha-D-Galp-(1->3)]-D-Galp Chemical compound O[C@H]1[C@H](O)[C@H](O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](O[C@@H]2[C@@H]([C@@H](O)[C@@H](O)[C@@H](CO)O2)O)[C@@H](O)[C@@H](CO)OC1O XNBZPOHDTUWNMW-OUUCXATCSA-N 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- 229960001230 asparagine Drugs 0.000 description 1
- 235000009582 asparagine Nutrition 0.000 description 1
- 125000000613 asparagine group Chemical group N[C@@H](CC(N)=O)C(=O)* 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000008827 biological function Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 125000001549 ceramide group Chemical group 0.000 description 1
- 239000011365 complex material Substances 0.000 description 1
- YSMODUONRAFBET-UHFFFAOYSA-N delta-DL-hydroxylysine Natural products NCC(O)CCC(N)C(O)=O YSMODUONRAFBET-UHFFFAOYSA-N 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 239000002158 endotoxin Substances 0.000 description 1
- YSMODUONRAFBET-UHNVWZDZSA-N erythro-5-hydroxy-L-lysine Chemical compound NC[C@H](O)CC[C@H](N)C(O)=O YSMODUONRAFBET-UHNVWZDZSA-N 0.000 description 1
- DQYBDCGIPTYXML-UHFFFAOYSA-N ethoxyethane;hydrate Chemical compound O.CCOCC DQYBDCGIPTYXML-UHFFFAOYSA-N 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- FJEKYHHLGZLYAT-FKUIBCNASA-N galp Chemical compound C([C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCSC)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(O)=O)C(=O)NCC(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(O)=O)C(=O)NCC(=O)N[C@@H](CC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC=1N=CNC=1)C(=O)N1[C@@H](CCC1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCC(N)=O)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CO)C(O)=O)[C@@H](C)CC)[C@@H](C)O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](NC(=O)[C@H]1N(CCC1)C(=O)CNC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)CNC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)CNC(=O)CNC(=O)[C@H](CCCNC(N)=N)NC(=O)CNC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@H](C)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](C)N)[C@@H](C)O)C(C)C)C1=CNC=N1 FJEKYHHLGZLYAT-FKUIBCNASA-N 0.000 description 1
- 238000002290 gas chromatography-mass spectrometry Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 230000035931 haemagglutination Effects 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 238000005984 hydrogenation reaction Methods 0.000 description 1
- QJHBJHUKURJDLG-UHFFFAOYSA-N hydroxy-L-lysine Natural products NCCCCC(NO)C(O)=O QJHBJHUKURJDLG-UHFFFAOYSA-N 0.000 description 1
- 230000008611 intercellular interaction Effects 0.000 description 1
- 230000000968 intestinal effect Effects 0.000 description 1
- GZQKNULLWNGMCW-PWQABINMSA-N lipid A (E. coli) Chemical compound O1[C@H](CO)[C@@H](OP(O)(O)=O)[C@H](OC(=O)C[C@@H](CCCCCCCCCCC)OC(=O)CCCCCCCCCCCCC)[C@@H](NC(=O)C[C@@H](CCCCCCCCCCC)OC(=O)CCCCCCCCCCC)[C@@H]1OC[C@@H]1[C@@H](O)[C@H](OC(=O)C[C@H](O)CCCCCCCCCCC)[C@@H](NC(=O)C[C@H](O)CCCCCCCCCCC)[C@@H](OP(O)(O)=O)O1 GZQKNULLWNGMCW-PWQABINMSA-N 0.000 description 1
- 229920006008 lipopolysaccharide Polymers 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 210000004400 mucous membrane Anatomy 0.000 description 1
- MBLBDJOUHNCFQT-UHFFFAOYSA-N n-(3,4,5,6-tetrahydroxy-1-oxohexan-2-yl)acetamide Chemical class CC(=O)NC(C=O)C(O)C(O)C(O)CO MBLBDJOUHNCFQT-UHFFFAOYSA-N 0.000 description 1
- 150000003019 phosphosphingolipids Chemical class 0.000 description 1
- 108020001775 protein parts Proteins 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
- 239000010935 stainless steel Substances 0.000 description 1
- 125000001424 substituent group Chemical group 0.000 description 1
- 231100000765 toxin Toxicity 0.000 description 1
- 238000005583 trifluoroacetylation reaction Methods 0.000 description 1
- 150000004043 trisaccharides Chemical class 0.000 description 1
- 210000003741 urothelium Anatomy 0.000 description 1
- 238000009834 vaporization Methods 0.000 description 1
- 230000008016 vaporization Effects 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H3/00—Compounds containing only hydrogen atoms and saccharide radicals having only carbon, hydrogen, and oxygen atoms
- C07H3/06—Oligosaccharides, i.e. having three to five saccharide radicals attached to each other by glycosidic linkages
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
- C07K16/34—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against blood group antigens
-
- 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
- G01N33/80—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving blood groups or blood types or red blood cells
Definitions
- the present invention relates to a process of isolating from different types of cell tissue carbohydrates mainly consisting of oligosaccharides with degradation of glycoconjugates present in the cell tissue, said glycoconjugate being a common term for glycoproteins, proteoglycanes and glycolipids.
- Cell tissue is a complex material consisting inter alia of so-called glycoconjugates, which are constituted by complex structures composed of carbohydrates, proteins and lipids.
- the three main types of glycoconjugates are glycoproteins, proteoglycanates and glycolipids.
- the carbohydrate chains are mostly N-glycosidically bound to asparagine or O-glycosidically to serine, treonine, hydroxylysine or hydr ⁇ xyproline.
- the carbohydrate chains of the proteoglycanes have similar bonds to the protein part.
- the carbohydrate part is O-glycosidically bound and is present as glycosyl ceramides in man and animals.
- the carbohydrate part of cell membrane glycoconjugates has been found to play an essential role in several biological functions including cell-cell interactions, as receptors for antibodies, enzymes, hormones, viruses and toxines.
- cell-cell interactions As receptors for antibodies, enzymes, hormones, viruses and toxines.
- the present invention has for its purpose to provide a process for releasing the carbohydrate part from the glycoconjugates in cell tissue while maintaining the carbohydrate chains in a more or less intact state. More in particular, the invention provides for a process for isolating the carbohydrate fraction from glycolipids of cell tissue by specifically cleaving the glycosidic bond between the carbohydrate part and the ceramide residue.
- the present invention is based on technique residing in treating the cell tissue with a reactant containing or consisting of trifluoroacetic acid anhydride.
- a reactant containing or consisting of trifluoroacetic acid anhydride By this treatment, which can be termed trifluoroacetolysis, the oligosaccharide chains are released from both glycoproteins and glycolipids with simultaneous degradation of proteins and lipids, whereby the oligosaccharides in the form of their trifluoroacetates can be separated and recovered.
- trifluoroacetic acid anhydride solely or trifluoroacetic acid anhydride in combination with trifluoroacetic acid, the latter constituting up to about 90 % by volume of the reactant mixture.
- a preferred reactant is one wherein trifluoroacetic acid anhydride and trifluoroacetic acid are present in about equal proportions based on volume.
- the treatment with the reactant is preferably carried out at an increased temperature and at an increased pressure.
- a preferred temperature range is 80-110°C, for example about 100°C.
- the overpressure in the latter case is about 4 atms.
- the cell tissue is constituted by erytrocyte membranes from which can be recovered oligosaccharides possessing blood group specificity.
- Such blood group specific oligosaccharides recovered in accordance with the technique of this invention can be coupled to carriers with antigenicity, and conjugates obtained thereby may then be used for immunisation for the purpose of obtaining specific antibodies.
- An alternative use of isolated oligosaccharides is coupling of said saccharides to carriers for the preparation of immunosorbent columns which can be used. for purification and isolation of specific antibodies.
- the specific antibodies made can be used in reactants for blood group determination and for other medicinal purposes.
- the reaction mixture obtained when treating the cell tissue with the reactant is suitably evaporized to dryness, and the residue vaporization is distributed between water and a water-immiscible solvent, the aqueous phase containing the carbohydrate fraction being recovered and the solvent phase containing inter alia protein residues and lipid residues being discarded.
- a water-immiscible solvent there may be used for example ether or methylene chloride.
- the erytrocyte membranes are suitably homogenized and suspended in water, whereafter lyophilisation is carried out before the treatment with the reactant.
- the oligosaccharides recovered by the process according to this invention may suitably be purified by gelchromatography and preparative paper chromatography. In this manner there may be obtained from blood of the blood group A the oligosaccharide: ⁇ -GalNAcp-(1-3)-Gal (A-tri) ⁇ -Galp-(1-4)- ⁇ -Galp-(1-4)-Glc or ⁇ -GalNAcp-(1-3)- ⁇ -Galp-(1-4)- ⁇ -Galp-(1-4)-Glc.
- oligosaccharides mentioned under A- and B-blood are obtained from AB-blood.
- the technique according to this invention namely release and recovery of oligosaccharides from erytrocyte membranes which show blood group specificity
- four main steps are part of the procedure, namely the following: a. Isolation of blood group specific oligosaccharides from erytrocyte membranes; b. Chemical modification of isolated oligosaccharide for coupling to an antigenic carrier or a carrier for the preparation of immunosorbent.
- d Purification of antibodies.
- EXAMPLES a Preparation of oligosaccharides. Erytrocytes from discarded blood of blood groups A, B, AB and O are lysed and the membranes are isolated by means of centrifugation. In this way about 10 grams of membranes are obtained from one liter of blood. The membranes are treated according to the process of Dodge, J.T., Mitchell, C, and Hanahan, D.J. (1963),
- reaction mixture is cooled and evaporized to dryness, there being obtained a dark to black-coloured residue.
- residue there is added methanol (700 ml), and the mixture is evaporised to dryness.
- the residue is then diluted with 50 % aqueous solution of acetic acid (1000 ml), and is allowed to star. ⁇ at room temperature for about l8 hours.
- the reaction mixture is filtered with a glass filter and evaporised to dryness.
- the residue obtained is distributed between water and diethyl ether.
- the ether phase is washed 4 times with water, and the combined wash solutions are washed with diethyl ether 4 times.
- the aqueous solution obtained is yellow-coloured and contains the released oligcsaccharides.
- the oligosaccharides obtained are in such form that the N-acetyl functions have been converted to N-trifluoroacetyl groups.
- the oligosaccharides obtained can be purified by gel chromatography followed by a preparative paper chromatography, and the purification procedure is continuously surveyed with gas chromatography mass spectrometry.
- the following oligosaccharides are obtained in the present example by treatment of blood originating from blood groups A, B, AB and O with varying blood groups within the P-system:
- Chemical modification of isolated oligosaccharides In order that the oligosaccharides shall be capable of coupling to carriers of different kinds they should be chemically modified. Such modification can be performed by means of known technique. Before the chemical modification it is suitable to de-N-trifluoroacetylate the oligosaccharides, which is suitably carried out in a basic environment at a pH of about 10-12 and at room temperatures for a period of time of about 20 hours. This results in the formation of amino groups, which can be N-acetylized to reform the N-acetyl functions.
- the aliphatic amino groups formed by the de-N-trifluoroacetylation can be used for coupling the modified oligosaccharide to different carrier materials.
- the modified oligosaccharides can be coupled to proteins, which can be constituted by albumen or edestine, and possibly to lipid A (from lipopolysaccharide) for the purpose of obtaining a high yield of M-antibodies.
- the conjugates obtained are then used for immunising animals, for example rabbit or horse, or for the production of antibodies by hybridom technique (Kohler, G. and Milstein, C, 1975, Nature (London) 256, pp. 495-497; Kohler G., and Milstein, C, 1976, Eur.J. Immunol. 6 pp. 511-519).
- the antisera thereby obtained are then serologically used in connection with known blood classification methods.
- d Purification of antibodies.
- Chemically modified blood group active oligosaccharides prepared according to the above are coupled to a suitable carrier material, for example Sephadex, and then sera from immunised animals are passed through an immunosorbent column formed by the treated Sephadex.
- Antibodies having specificity vis-a-vis coupled oligosaccharide adheres by reaction with same, whereas materials of no interest are washed away.
- the specific antibodies adhering to the column by reaction with coupled oligosaccharides are then desorbed, and antibody solutions of unambiguously defined specificity are obtained.
- Purified specific antibodies ar ⁇ then tested for specificity for control and may then be used for blood group determination by hemagglutination technique.
- the present invention is not delimited to the exemplary application for preparation of blood group active oligosaccharides from erytrocyte membranes as described above.
- the invention is applicable to all types of cell tissue for recovery and isolation of oligosaccharides for different purposes.
- epithelium tissue intestinal mucous membrane, urinary tract epithelium, lung epithelium etc.
- isolated oligosaccharides can be of diagnostic importance.
- GalNAcp 2-acetamido-2-deoxy-D-galactopyranosyl
- Gal D-galactose
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Immunology (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Biochemistry (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Biotechnology (AREA)
- Genetics & Genomics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Food Science & Technology (AREA)
- Pharmacology & Pharmacy (AREA)
- Biophysics (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Cell Biology (AREA)
- Microbiology (AREA)
- Veterinary Medicine (AREA)
- Epidemiology (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Saccharide Compounds (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
Procede d'isolation de la fraction hydrates de carbone (oligosaccharides) a partir de tissus cellulaires avec degradation des glycoconjugues, le tissu cellulaire etant traite avec un reactif contenant ou se composant d'anhydride d'acide trifluoroacetique, les hydrates de carbone etant liberes a la suite de leur conversion en trifluoroacetates, qui sont facultativement detrifluoroacetyles. En partant de membranes erythrocytes, des oligosaccharides possedant une specificite de groupes sanguins peuvent etre recuperes, a partir de tels oligosaccharides et sont utiles pour la determination du groupe sanguin.Method for isolating the carbohydrate fraction (oligosaccharides) from cell tissues with degradation of the glycoconjugates, the cell tissue being treated with a reagent containing or consisting of trifluoroacetic acid anhydride, the carbohydrates being released to following their conversion to trifluoroacetates, which are optionally detrifluoroacetylated. Starting from erythrocyte membranes, oligosaccharides having blood group specificity can be recovered from such oligosaccharides and are useful for blood group determination.
Description
Claims
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
SE8000658A SE8000658L (en) | 1980-01-28 | 1980-01-28 | ISOLATION OF CARBON HYDRATES FROM CELL TISSUE |
SE8000658 | 1980-01-28 |
Publications (1)
Publication Number | Publication Date |
---|---|
EP0051593A1 true EP0051593A1 (en) | 1982-05-19 |
Family
ID=20340092
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
EP81900324A Withdrawn EP0051593A1 (en) | 1980-01-28 | 1981-01-28 | Isolation of carbohydrates from cell tissue |
Country Status (6)
Country | Link |
---|---|
EP (1) | EP0051593A1 (en) |
JP (1) | JPS57500022A (en) |
DK (1) | DK427581A (en) |
IT (1) | IT1141962B (en) |
SE (1) | SE8000658L (en) |
WO (1) | WO1981002104A1 (en) |
Families Citing this family (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
GB2117789B (en) * | 1982-03-29 | 1985-08-29 | East Anglian Regional Health | Abo blood grouping reagent |
SE8301609D0 (en) * | 1983-03-23 | 1983-03-23 | Svenska Sockerfabriks Ab | ASSOCIATION AND COMPOSITION OF THERAPEUTIC OR DIAGNOSTIC APPLICATION AS PROCEDURES FOR THERAPEUTIC TREATMENT |
SE8304006D0 (en) * | 1983-07-15 | 1983-07-15 | Karlsson Karl Anders | ASSOCIATION AND COMPOSITION OF THERAPEUTIC OR DIAGNOSTIC USE AND PROCEDURE FOR THERAPEUTIC TREATMENT |
FR2553518B1 (en) * | 1983-10-13 | 1986-04-18 | Choay Sa | NEW CONJUGATES DEVELOPED BY FIXING A LIGAND ON AN INSOLUBLE SUPPORT, THEIR PREPARATION AND THEIR BIOLOGICAL APPLICATIONS |
-
1980
- 1980-01-28 SE SE8000658A patent/SE8000658L/en not_active Application Discontinuation
-
1981
- 1981-01-28 EP EP81900324A patent/EP0051593A1/en not_active Withdrawn
- 1981-01-28 WO PCT/SE1981/000021 patent/WO1981002104A1/en active IP Right Grant
- 1981-01-28 JP JP56500550A patent/JPS57500022A/ja active Pending
- 1981-01-28 IT IT19379/81A patent/IT1141962B/en active
- 1981-09-25 DK DK427581A patent/DK427581A/en not_active Application Discontinuation
Non-Patent Citations (1)
Title |
---|
See references of WO8102104A1 * |
Also Published As
Publication number | Publication date |
---|---|
IT8119379A0 (en) | 1981-01-28 |
WO1981002104A1 (en) | 1981-08-06 |
IT1141962B (en) | 1986-10-08 |
JPS57500022A (en) | 1982-01-07 |
DK427581A (en) | 1981-09-25 |
SE8000658L (en) | 1981-07-29 |
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